We present a large-scale molecular phylogeny of the ants (Hymenoptera: Formicidae), based on 4.5 kilobases of sequence data from six gene regions extracted from 139 of the 288 described extant genera, representing 19 of the 20 subfamilies. All but two subfamilies are recovered as monophyletic. Divergence time estimates calibrated by minimum age constraints from 43 fossils indicate that most of the subfamilies representing extant ants arose much earlier than previously proposed but only began to diversify during the Late Cretaceous to Early Eocene. This period also witnessed the rise of angiosperms and most herbivorous insects.

Summary: The generalized mixed Yule‐coalescent (GMYC) model has become one of the most popular approaches for species delimitation based on single‐locus data, and it is widely used in biodiversity assessments and phylogenetic community ecology. We here examine an array of factors affecting GMYC resolution (tree reconstruction method, taxon sampling coverage/taxon richness and geographic sampling intensity/geographic scale). We test GMYC performance based on empirical data (DNA barcoding of the Romanian butterflies) on a solid taxonomic framework (i.e. all species are thought to be described and can be determined with independent sources of evidence). The data set is comprehensive (176 species), and intensely and homogeneously sampled (1303 samples representing the main populations of butterflies in this country). Taxonomy was assessed based on morphology, including linear and geometric morphometry when needed. The number of GMYC entities obtained constantly exceeds the total number of morphospecies in the data set. We show that c. 80% of the species studied are recognized as entities by GMYC. Interestingly, we show that this percentage is practically the maximum that a single‐threshold method can provide for this data set. Thus, the c. 20% of failures are attributable to intrinsic properties of the COI polymorphism: overlap in inter‐ and intraspecific divergences and non‐monophyly of the species likely because of introgression or lack of independent lineage sorting. Our results demonstrate that this method is remarkably stable under a wide array of circumstances, including most phylogenetic reconstruction methods, high singleton presence (up to 95%), taxon richness (above five species) and the presence of gaps in intraspecific sampling coverage (removal of intermediate haplotypes). Hence, the method is useful to designate an optimal divergence threshold in an objective manner and to pinpoint potential cryptic species that are worth being studied in detail. However, the existence of a substantial percentage of species wrongly delimited indicates that GMYC cannot be used as sufficient evidence for evaluating the specific status of particular cases without additional data. Finally, we provide a set of guidelines to maximize efficiency in GMYC analyses and discuss the range of studies that can take advantage of the method.

The proliferation of DNA data is revolutionizing all fields of systematic research. DNA barcode sequences, now available for millions of specimens and several hundred thousand species, are increasingly used in algorithmic species delimitations. This is complicated by occasional incongruences between species and gene genealogies, as indicated by situations where conspecific individuals do not form a monophyletic cluster in a gene tree. In two previous reviews, non-monophyly has been reported as being common in mitochondrial DNA gene trees. We developed a novel web service “Monophylizer” to detect non-monophyly in phylogenetic trees and used it to ascertain the incidence of species non-monophyly in COI (a.k.a. cox1) barcode sequence data from 4977 species and 41,583 specimens of European Lepidoptera, the largest data set of DNA barcodes analyzed from this regard. Particular attention was paid to accurate species identification to ensure data integrity. We investigated the effects of tree-building method, sampling effort, and other methodological issues, all of which can influence estimates of non-monophyly. We found a 12% incidence of non-monophyly, a value significantly lower than that observed in previous studies. Neighbor joining (NJ) and maximum likelihood (ML) methods yielded almost equal numbers of non-monophyletic species, but 24.1% of these cases of non-monophyly were only found by one of these methods. Non-monophyletic species tend to show either low genetic distances to their nearest neighbors or exceptionally high levels of intraspecific variability. Cases of polyphyly in COI trees arising as a result of deep intraspecific divergence are negligible, as the detected cases reflected misidentifications or methodological errors. Taking into consideration variation in sampling effort, we estimate that the true incidence of non-monophyly is ∼23%, but with operational factors still being included. Within the operational factors, we separately assessed the frequency of taxonomic limitations (presence of overlooked cryptic and oversplit species) and identification uncertainties. We observed that operational factors are potentially present in more than half (58.6%) of the detected cases of non-monophyly. Furthermore, we observed that in about 20% of non-monophyletic species and entangled species, the lineages involved are either allopatric or parapatric—conditions where species delimitation is inherently subjective and particularly dependent on the species concept that has been adopted. These observations suggest that species-level non-monophyly in COI gene trees is less common than previously supposed, with many cases reflecting misidentifications, the subjectivity of species delimitation or other operational factors.

Phylogenetic relationships among major clades of butterflies and skippers have long been controversial, with no general consensus even today. Such lack of resolution is a substantial impediment to using the otherwise well studied butterflies as a model group in biology. Here we report the results of a combined analysis of DNA sequences from three genes and a morphological data matrix for 57 taxa (3258 characters, 1290 parsimony informative) representing all major lineages from the three putative butterfly super-families (Hedyloidea, Hesperioidea and Papilionoidea), plus out-groups representing other ditrysian Lepidoptera families. Recently, the utility of morphological data as a source of phylogenetic evidence has been debated. We present the first well supported phylogenetic hypothesis for the butterflies and skippers based on a total-evidence analysis of both traditional morphological characters and new molecular characters from three gene regions (COI,EF-1αandwingless). All four data partitions show substantial hidden support for the deeper nodes, which emerges only in a combined analysis in which the addition of morphological data plays a crucial role. With the exception of Nymphalidae, the traditionally recognized families are found to be strongly supported monophyletic clades with the following relationships: (Hesperiidae+(Papilionidae+(Pieridae+(Nymphalidae+(Lycaenidae+Riodinidae))))). Nymphalidae is recovered as a monophyletic clade but this clade does not have strong support. Lycaenidae and Riodinidae are sister groups with strong support and we suggest that the latter be given family rank. The position of Pieridae as the sister taxon to nymphalids, lycaenids and riodinids is supported by morphology and theEF-1αdata but conflicted by theCOIandwinglessdata. Hedylidae are more likely to be related to butterflies and skippers than geometrid moths and appear to be the sister group to Papilionoidea+Hesperioidea.

Butterflies are a diverse and charismatic insect group that are thought to have evolved with plants and dispersed throughout the world in response to key geological events. However, these hypotheses have not been extensively tested because a comprehensive phylogenetic framework and datasets for butterfly larval hosts and global distributions are lacking. We sequenced 391 genes from nearly 2,300 butterfly species, sampled from 90 countries and 28 specimen collections, to reconstruct a new phylogenomic tree of butterflies representing 92% of all genera. Our phylogeny has strong support for nearly all nodes and demonstrates that at least 36 butterfly tribes require reclassification. Divergence time analyses imply an origin ~100 million years ago for butterflies and indicate that all but one family were present before the K/Pg extinction event. We aggregated larval host datasets and global distribution records and found that butterflies are likely to have first fed on Fabaceae and originated in what is now the Americas. Soon after the Cretaceous Thermal Maximum, butterflies crossed Beringia and diversified in the Palaeotropics. Our results also reveal that most butterfly species are specialists that feed on only one larval host plant family. However, generalist butterflies that consume two or more plant families usually feed on closely related plants.