Jorge Llorente-Bousquets

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.

UV Photoreceptors and UV-Yellow Wing Pigments in Heliconius Butterflies Allow a Color Signal to Serve both Mimicry and Intraspecific Communication

Mimetic wing coloration evolves in butterflies in the context of predator confusion. Unless butterfly eyes have adaptations for discriminating mimetic color variation, mimicry also carries a risk of confusion for the butterflies themselves. Heliconius butterfly eyes, which express recently duplicated ultraviolet (UV) opsins, have such an adaptation. To examine bird and butterfly color vision as sources of selection on butterfly coloration, we studied yellow wing pigmentation in the tribe Heliconiini. We confirmed, using reflectance and mass spectrometry, that only Heliconius use 3-hydroxy-DL-kynurenine (3-OHK), which looks yellow to humans but reflects both UV- and long-wavelength light, whereas butterflies in related genera have chemically unknown yellow pigments mostly lacking UV reflectance. Modeling of these color signals reveals that the two UV photoreceptors of Heliconius are better suited to separating 3-OHK from non-3-OHK spectra compared with the photoreceptors of related genera or birds. The co-occurrence of potentially enhanced UV vision and a UV-reflecting yellow wing pigment could allow unpalatable Heliconius private intraspecific communication in the presence of mimics. Our results are the best available evidence for the correlated evolution of a color signal and color vision. They also suggest that predator visual systems are error prone in the context of mimicry.

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.

Chorionic Morphology in the Coliadinae Subfamily (Lepidoptera: Pieridae)

Abstract. We described and compared the chorion morphology of 22 species in 12 genera of the subfamily Coliadinae, collected from 80 females in Mexico, Colombia, and Ecuador. Their characterization and ilustration include the main structural features of the eggs: colors, shapes, sizes, and diversity of ornamentation or reticula; possibly from the most widespread in Pieridae (polygon grid) to the derivatives Coliadinae (with the formation of axes and ribs relatively reduced number). We describe for the first time microreticulum associated with the macroreticulum.

Morfología del Corion en Leptophobia (Lepidoptera: Pieridae) e Importancia Taxonómica

Resumen. Se describe y compara el corion de siete especies del género Leptophobia Butler, 1870: L. eleusis eleusis (Lucas, 1852); L. tovaria maruga Fruhstorfer, 1907; L. cinerea cinerea (Hewitson, 1867); L. caesia caesia (Lucas, 1852); L. eleone luca Fruhstorfer, 1907; L. philoma subargentea (Hewitson, 1870); y L. aripa elodia (Boisduval, 1836). Su descripción e ilustración considera los siguientes rasgos estructurales: número y disposición de ejes (cortos y largos), cuantificación de costillas, descripción de la trama y la distinción de protuberancias en la región perimicropilar. También se describe el color del huevo y su tamaño (longitud y su proporción con el diámetro máximo). Se tipifican tres formas de huevo: elipsoidal oblonga, barrilete, y obovada. Se hacen comparaciones entre los caracteres presentes en las diferentes especies; se comentan implicaciones taxonómicas y filogenéticas. Los caracteres del corion están correlacionados con los caracteres alares y el ambiente de vuelo de los imagos, esto lleva a reconocer dos subgrupos de especies en el género (hipótesis de Klots).

Chorion Morphology in the Genus Ascia and Ganyra and Its Comparison with other Proximate Genera to Pierinae (Lepidoptera: Pieridae)

Abstract. We describe and compare the chorion morphology of three species of the genus Ascia and Ganyra. Their characterization and illustration include the main structural features of the egg: number and arrangement of longitudinal axes (short and long), and quantification of ribs or cells, protuberances, projections, and reminiscent ribs; the color and size are also described. We typify three egg models: ellipsoidal with protuberances exhibiting a grid of pores in Ascia, elongated or oblong ellipsoidal with laminar projections unciforms in Ganyra. Comparisons are made between the chorion features of different species and related genera (Glutophrissa and Leptophobia). We discuss their phylogenetic and taxonomic implications.

Morphological Comparison of the Chorion of the Genera Pieriballia, Itaballia, and Perrhybris (Lepidoptera: Pieridae: Pierinae), and their Phylogenetic Implications

Abstract. We describe and compare the egg chorion of five species of three genera of Pierinae: Pieriballia, Itaballia, and Perrhybris. Their characterizations and illustrations include the main structural features: number and arrangement of longitudinal axes (short and long), and quantification of ribs or cells, projections and vestiges or reminiscent ribs. Also, we describe the color and size of the egg. We typify two egg models: elongated ellipsoidal or fusiform and oblong ellipsoidal, both with projections in the apical area, around the micropyle. Comparisons are made between features present in different species and genera. We discuss their phylogenetic and taxonomic implications.

Sexual Dimorphism and Retinal Mosaic Diversification Following the Evolution of a Violet Receptor in Butterflies.

Numerous animal lineages have expanded and diversified the opsin-based photoreceptors in their eyes underlying color vision behavior. However, the selective pressures giving rise to new photoreceptors and their spectral tuning remain mostly obscure. Previously, we identified a violet receptor (UV2) that is the result of a UV opsin gene duplication specific to Heliconius butterflies. At the same time the violet receptor evolved, Heliconius evolved UV-yellow coloration on their wings, due to the pigment 3-hydroxykynurenine (3-OHK) and the nanostructure architecture of the scale cells. In order to better understand the selective pressures giving rise to the violet receptor, we characterized opsin expression patterns using immunostaining (14 species) and RNA-Seq (18 species), and reconstructed evolutionary histories of visual traits in five major lineages within Heliconius and one species from the genus Eueides. Opsin expression patterns are hyperdiverse within Heliconius. We identified six unique retinal mosaics and three distinct forms of sexual dimorphism based on ommatidial types within the genus Heliconius. Additionally, phylogenetic analysis revealed independent losses of opsin expression, pseudogenization events, and relaxation of selection on UVRh2 in one lineage. Despite this diversity, the newly evolved violet receptor is retained across most species and sexes surveyed. Discriminability modeling of behaviorally preferred 3-OHK yellow wing coloration suggests that the violet receptor may facilitate Heliconius color vision in the context of conspecific recognition. Our observations give insights into the selective pressures underlying the origins of new visual receptors.

Morfología Coriónica de Once Especies de Biblidinae1 de México y Colombia: un Examen de Predicciones

Resumen. Se complementa la descripción e ilustración relativamente detallada del corion en la subfamilia Biblidinae, al agregarse 11 especies distintas a las presentadas recientemente, en los géneros Hamadryas, Marpesia, Eunica, Catonephele, Myscelia, Asterope, Epiphile, Callicore, y Cyclogramma. Los ejemplares provienen de México y Colombia. El estudio se realizó a partir de una muestra de 35 hembras, de las cuales 15 tuvieron huevos maduros. Se caracterizaron los principales rasgos estructurales del corion: color, forma, tamaño y relieve: diferenciación perimicropilar y formación de ejes y costillas. Se hace una distinción de rasgos especializados en las zonas apical y ecuatorial, así como el grado de engrosamiento de los ejes. Se siguen los tipos de huevos reconocidos recientemente, así mismo la nomenclatura de las descripciones. Los resultados confirman los tipos de huevo y arquitectura propias de cada taxón supragenérico. Además, apoyan la propuesta de emplear al corion como carácter taxonómico, al encontrarse características singulares de cada género y especie. Cabe añadir que una vez más se sustenta la posibilidad de resurgir la subtribu Eunicina de Reuter, al escindir a Eunica de la subtribu Epicaliina. La similitud coriónica entre Cyclogramma y Diaethria apoya la propuesta de Lamas de considerar sinónimos a estos géneros.

Reply to Nozawa et al.: Complementary statistical methods support positive selection of a duplicated UV opsin gene in Heliconius

Statistical methods used to test for positive selection have a long history and continue to evolve (1–4). In their letter, Nozawa et al. (5) question our use of the branch-site method in our recent paper in PNAS (6). As experimental biologists, we welcome all methods that facilitate the detection of interesting parts of the genome for functional exploration. In this case, a statistically significant result using the branch-site method combined with structural modeling and the identification of a few biochemically relevant substitutions provided us with an incentive for the in vivo physiological characterization of the UV-sensitive rhodopsins in Heliconius. Had we not had that first hint from sequence data alone, it is unlikely that we would have made that functional discovery, which makes the eyes of Heliconius unique compared with all other studied butterflies.