Andrew D. Warren

Positive selection of a duplicated UV-sensitive visual pigment coincides with wing pigment evolution in Heliconius butterflies.

The butterfly Heliconius erato can see from the UV to the red part of the light spectrum with color vision proven from 440 to 640 nm. Its eye is known to contain three visual pigments, rhodopsins, produced by an 11-cis-3-hydroxyretinal chromophore together with long wavelength (LWRh), blue (BRh) and UV (UVRh1) opsins. We now find that H. erato has a second UV opsin mRNA (UVRh2)—a previously undescribed duplication of this gene among Lepidoptera. To investigate its evolutionary origin, we screened eye cDNAs from 14 butterfly species in the subfamily Heliconiinae and found both copies only among Heliconius. Phylogeny-based tests of selection indicate positive selection of UVRh2 following duplication, and some of the positively selected sites correspond to vertebrate visual pigment spectral tuning residues. Epi-microspectrophotometry reveals two UV-absorbing rhodopsins in the H. erato eye with λmax = 355 nm and 398 nm. Along with the additional UV opsin, Heliconius have also evolved 3-hydroxy-DL-kynurenine (3-OHK)-based yellow wing pigments not found in close relatives. Visual models of how butterflies perceive wing color variation indicate this has resulted in an expansion of the number of distinguishable yellow colors on Heliconius wings. Functional diversification of the UV-sensitive visual pigments may help explain why the yellow wing pigments of Heliconius are so colorful in the UV range compared to the yellow pigments of close relatives lacking the UV opsin duplicate.

Revised classification of the family Hesperiidae (Lepidoptera: Hesperioidea) based on combined molecular and morphological data

Abstract We propose a revised higher classification for the genera of Hesperiidae (skipper butterflies) of the world. We have augmented our published DNA data matrix with 49 morphological characters in order to infer relationships for taxa not sampled in the molecular study. We use the results of a combined analysis to identify morphological synapomorphies of the suprageneric clades of Hesperiidae, and to hypothesize a phylogenetic classification of the world’s genera of Hesperiidae, the first of its kind for this diverse group. Monophyly of the family Hesperiidae is strongly supported, as are some of the traditionally recognized subfamilies. The results presented here largely corroborate those of our molecular study, but differ in several details. The Australian endemic Euschemon rafflesia is given subfamily status, as is Eudaminae. We recognize seven subfamilies of Hesperiidae: Coeliadinae, Euschemoninae (confirmed status), Eudaminae (new status), Pyrginae, Heteropterinae (confirmed status), Trapezitinae and Hesperiinae. We treat Pyrrhopygini, Tagiadini, Celaenorrhinini, Carcharodini, Achlyodidini, Erynnini and Pyrgini as tribes of Pyrginae. Circumscriptions of Achlyodidini and Pyrgini require further elucidation. Tribes of Hesperiinae include Aeromachini, Baorini, Taractrocerini, Thymelicini, Calpodini (reinstated status), Anthoptini (new tribe), Moncini and Hesperiini. The tribal placement of many Old World hesperiine genera remains ambiguous.

Phylogenetic relationships of subfamilies and circumscription of tribes in the family Hesperiidae (Lepidoptera: Hesperioidea)

A comprehensive tribal‐level classification for the world’s subfamilies of Hesperiidae, the skipper butterflies, is proposed for the first time. Phylogenetic relationships between tribes and subfamilies are inferred using DNA sequence data from three gene regions (cytochrome oxidase subunit I‐subunit II, elongation factor‐1α and wingless). Monophyly of the family is strongly supported, as are some of the traditionally recognized subfamilies, with the following relationships: (Coeliadinae + (“Pyrginae” + (Heteropterinae + (Trapezitinae + Hesperiinae)))). The subfamily Pyrginae of contemporary authors was recovered as a paraphyletic grade of taxa. The formerly recognized subfamily Pyrrhopyginae, although monophyletic, is downgraded to a tribe of the “Pyrginae”. The former subfamily Megathyminae is an infra‐tribal group of the Hesperiinae. The Australian endemic Euschemon rafflesia is a hesperiid, possibly related to “Pyrginae” (Eudamini). Most of the traditionally recognized groups and subgroups of genera currently employed to partition the subfamilies of the Hesperiidae are not monophyletic. We recognize eight pyrgine and six hesperiine tribes, including the new tribe Moncini.

Seasonality and phenology of the butterflies (Lepidoptera : Papilionoidea and Hesperioidea) of Mexico's Calakmul Region

Abstract The phenology of butterflies was analyzed in the Calakmul Region (CR) in the state of Campeche, México, over the course of 3 years. Altogether, 60,662 individuals were recorded, consisting of 359 species in 207 genera, 18 subfamilies, 5 families, and 2 superfamilies. Greatest species diversity was recorded during Oct and Nov. Monthly fluctuation in diversity was defined by rare species. Hesperiidae (135 species) and Nymphalidae (111 species) were the most diverse families, and showed the greatest variation with respect to distribution of species richness throughout the year. Papilionidae showed the greatest species richness during the dry season. Pieridae, Nymphalidae, and Lycaenidae showed peaks of greatest species richness and relative abundance during the rainy season. Results were compared to faunal studies of the Sierra de Atoyac de Álvarez, in the state of Guerrero, and of the Sierra de Manantlán, in Jalisco and Colima. Important similarities were observed among phenological patterns in the butterfly fauna of the 3 regions, especially between CR and Manantlán. The phenology of species with greater relative abundance was analyzed in relation to wingspan as a parameter of adult size. The small and medium-sized groups, taken together, showed variations in species richness. An analysis of species seasonality was conducted with NMDS, ANOSIM and SIMPER, in the program PRIMER 4.0. Differences among the composition of butterfly communities with respect to the seasons were found.

Speciation in Cloudless Sulphurs Gleaned from Complete Genomes

For 200 years, zoologists have relied on phenotypes to learn about the evolution of animals. A glance at the genotype, even through several gene markers, revolutionized our understanding of animal phylogeny. Recent advances in sequencing techniques allow researchers to study speciation mechanisms and the link between genotype and phenotype using complete genomes. We sequenced and assembled a complete genome of the Cloudless Sulphur (Phoebis sennae) from a single wild-caught specimen. This genome was used as reference to compare genomes of six specimens, three from the eastern populations (Oklahoma and north Texas), referred to as a subspecies Phoebis sennae eubule, and three from the southwestern populations (south Texas) known as a subspecies Phoebis sennae marcellina. While the two subspecies differ only subtly in phenotype and mitochondrial DNA, comparison of their complete genomes revealed consistent and significant differences, which are more prominent than those between tiger swallowtails Pterourus canadensis and Pterourus glaucus. The two sulphur taxa differed in histone methylation regulators, chromatin-associated proteins, circadian clock, and early development proteins. Despite being well separated on the whole-genome level, the two taxa show introgression, with gene flow mainly from P. s. marcellina to P. s. eubule. Functional analysis of introgressed genes reveals enrichment in transmembrane transporters. Many transporters are responsible for nutrient uptake, and their introgression may be of selective advantage for caterpillars to feed on more diverse food resources. Phylogenetically, complete genomes place family Pieridae away from Papilionidae, which is consistent with previous analyses based on several gene markers.

Hostplant change and paleoclimatic events explain diversification shifts in skipper butterflies (Family: Hesperiidae)

BackgroundSkippers (Family: Hesperiidae) are a large group of butterflies with ca. 4000 species under 567 genera. The lack of a time-calibrated higher-level phylogeny of the group has precluded understanding of its evolutionary past. We here use a 10-gene dataset to reconstruct the most comprehensive time-calibrated phylogeny of the group, and explore factors that affected the diversification of these butterflies.ResultsAncestral state reconstructions show that the early hesperiid lineages utilized dicots as larval hostplants. The ability to feed on monocots evolved once at the K-Pg boundary (ca. 65 million years ago (Mya)), and allowed monocot-feeders to diversify much faster on average than dicot-feeders. The increased diversification rate of the monocot-feeding clade is specifically attributed to rate shifts in two of its descendant lineages. The first rate shift, a four-fold increase compared to background rates, happened ca. 50 Mya, soon after the Paleocene-Eocene thermal maximum, in a lineage of the subfamily Hesperiinae that mostly fed on forest monocots. The second rate shift happened ca. 40 Mya in a grass-feeding lineage of Hesperiinae when open-habitat grasslands appeared in the Neotropics owing to gradual cooling of the atmospheric temperature.ConclusionsThe evolution of monocot feeding strongly influenced diversification of skippers. We hypothesize that although monocot feeding was an intrinsic trait that allowed exploration of novel niches, the lack of extensive availability of monocots comprised an extrinsic limitation for niche exploration. The shifts in diversification rate coincided with paleoclimatic events during which grasses and forest monocots were diversified.

Ten genes and two topologies: an exploration of higher relationships in skipper butterflies (Hesperiidae)

Despite multiple attempts to infer the higher-level phylogenetic relationships of skipper butterflies (Family Hesperiidae), uncertainties in the deep clade relationships persist. The most recent phylogenetic analysis included fewer than 30% of known genera and data from three gene markers. Here we reconstruct the higher-level relationships with a rich sampling of ten nuclear and mitochondrial markers (7,726 bp) from 270 genera and find two distinct but equally plausible topologies among subfamilies at the base of the tree. In one set of analyses, the nuclear markers suggest two contrasting topologies, one of which is supported by the mitochondrial dataset. However, another set of analyses suggests mito-nuclear conflict as the reason for topological incongruence. Neither topology is strongly supported, and we conclude that there is insufficient phylogenetic evidence in the molecular dataset to resolve these relationships. Nevertheless, taking morphological characters into consideration, we suggest that one of the topologies is more likely.

The Butterflies of Barro Colorado Island, Panama: Local Extinction since the 1930s

Few data are available about the regional or local extinction of tropical butterfly species. When confirmed, local extinction was often due to the loss of host-plant species. We used published lists and recent monitoring programs to evaluate changes in butterfly composition on Barro Colorado Island (BCI, Panama) between an old (1923–1943) and a recent (1993–2013) period. Although 601 butterfly species have been recorded from BCI during the 1923–2013 period, we estimate that 390 species are currently breeding on the island, including 34 cryptic species, currently only known by their DNA Barcode Index Number. Twenty-three butterfly species that were considered abundant during the old period could not be collected during the recent period, despite a much higher sampling effort in recent times. We consider these species locally extinct from BCI and they conservatively represent 6% of the estimated local pool of resident species. Extinct species represent distant phylogenetic branches and several families. The butterfly traits most likely to influence the probability of extinction were host growth form, wing size and host specificity, independently of the phylogenetic relationships among butterfly species. On BCI, most likely candidates for extinction were small hesperiids feeding on herbs (35% of extinct species). However, contrary to our working hypothesis, extinction of these species on BCI cannot be attributed to loss of host plants. In most cases these host plants remain extant, but they probably subsist at lower or more fragmented densities. Coupled with low dispersal power, this reduced availability of host plants has probably caused the local extinction of some butterfly species. Many more bird than butterfly species have been lost from BCI recently, confirming that small preserves may be far more effective at conserving invertebrates than vertebrates and, therefore, should not necessarily be neglected from a conservation viewpoint.

The evolutionary history of Boloria (Lepidoptera: Nymphalidae): phylogeny, zoogeography and larval–foodplant relationships

Boloria is restricted to cooler regions of the northern hemisphere, and represents a model for understanding recent diversification of phytophagous insects in the Holarctic region. We infer the phylogeny of the genus based on characters from adult morphology and three genes (COI, EF1-α and wingless). We revise the subgenus level classification, and divide the genus into three subgenera, corroborating earlier works. The largest subgenus, Clossiana, is divided into nine informal species groups. A relaxed Bayesian clock analysis combined with a dispersal-vicariance analysis shows that the genus originated and first diversified in the Central Palaearctic region in the Middle Eocene. This region has remained important throughout the evolution of the genus, but independent diversifications also occurred in the Nearctic region. At least nine independent dispersal events between the Palaearctic and Nearctic regions are needed to explain the current distribution of the genus, whereas vicariance appears to have played only a minor role. An analysis of the evolution of larval–foodplant associations shows that although monophagous Violaceae-feeding is the original feeding strategy within Boloria, numerous host-shifts or broadenings of diet have happened. This inherent plasticity in host choice has likely been an important factor in the diversification in alpine and arctic environments by Boloria.

Investigating Concordance among Genetic Data, Subspecies Circumscriptions and Hostplant Use in the Nymphalid Butterfly Polygonia faunus

Subspecies are commonly used taxonomic units to formally describe intraspecific geographic variation in morphological traits. However, the concept of subspecies is not clearly defined, and there is little agreement about what they represent in terms of evolutionary units, and whether they can be used as reliably useful units in conservation, evolutionary theory and taxonomy. We here investigate whether the morphologically well-characterized subspecies in the North American butterfly Polygonia faunus are supported by genetic data from mitochondrial sequences and eight microsatellite loci. We also investigate the phylogeographic structure of P. faunus and test whether similarities in host-plant use among populations are related to genetic similarity. Neither the nuclear nor the mitochondrial data corroborated subspecies groupings. We found three well defined genetic clusters corresponding to California, Arizona and (New Mexico+Colorado). There was little structuring among the remaining populations, probably due to gene flow across populations. We found no support for the hypothesis that similarities in host use are related to genetic proximity. The results indicate that the species underwent a recent rapid expansion, probably from two glacial refugia in western North America. The mitochondrial haplotype network indicates at least two independent expansion phases into eastern North America. Our results clearly demonstrate that subspecies in P. faunus do not conform to the structuring of genetic variation. More studies on insects and other invertebrates are needed to better understand the scope of this phenomenon. The results of this study will be crucial in designing further experiments to understand the evolution of hostplant utilization in this species.