Not all apparently gynandromorphic butterflies are gynandrous: The case of Polyommatus icarus and its relatives (Lepidoptera: Lycaenidae).

Beside the more than two thousand normal specimens of Polyommatus icarus (Rottemburg, 1775) yielded by rearing experiments, there was one perfectly bilateral dichromatic individual first considered to be gynandrous. On the basis of analysing genitalia traits, wing surface covering scale micromorphology, and the spectral characteristics of the blue colour generated by the cover scales, the gender of the specimen has been identified as female. This exemplar was investigated in comparison with gynandrous specimens from the collections of the Hungarian Natural History Museum exhibiting various degrees of intermixing of blue and brown coloration. Focus stacking microscopy for detailed scale morphology and UV-visible reflectance spectroscopy was used for the characterization of the optical properties. Inspecting literature references and the Lycaenidae collection of the museum, further examples have been found for female bilateral dichromatism in the closely related polyommatine lycaenid species Lysandra bellargus (Rottemburg, 1775) and Lysandra coridon (Poda, 1761) what suggests that polyommatine female dichromaticity may be displayed by the manner of bilaterality and mosaicism, phenomena hitherto solely connected to gynandromorphy.

Breeding Polyommatus icarus Serves as a Large-Scale and Environmentally Friendly Source of Precisely Tuned Photonic Nanoarchitectures.

The colour of the butterfly wing serves as an important sexual and species-specific signal. Some species produce structural colouration by developing wing scales with photonic nanoarchitectures. These nanostructures are highly conservative, allowing only a ±10 nm peak wavelength deviation in the reflectance spectra of the blue structural colour in natural Common Blue (Polyommatus icarus) populations. They are promising templates of future artificial photonic materials and can be used in potential applications, too. In this work, we present methodology and infrastructure for breeding laboratory populations of Common Blue as a cost-effective and environmentally friendly source of nanostructures. Our technology enables the production of approximately 7500 wing samples, equivalent to 0.5-1 m2 of photonic nanoarchitecture surface within a year in a single custom-made insectarium. To ascertain the reliability of this method, we compared reflectance properties between different populations from distant geographic locations. We also provide genetic background of these populations using microsatellite genotyping. The laboratory population showed genetic erosion, but even after four generations of inbreeding, only minimal shifts in the structural colouration were observed, indicating that wild Common Blue populations may be a reliable source of raw material for photonic surfaces.

Investigating the Effect of Reflectance Tuning on Photocatalytic Dye Degradation with Biotemplated ZnO Photonic Nanoarchitectures Based on Morpho Butterfly Wings.

Photonic nanoarchitectures of butterfly wings can serve as biotemplates to prepare semiconductor thin films of ZnO by atomic layer deposition. The resulting biotemplated ZnO nanoarchitecture preserves the structural and optical properties of the natural system, while it will also have the features of the functional material. The ZnO-coated wings can be used directly in heterogeneous photocatalysis to decompose pollutants dissolved in water upon visible light illumination. We used the photonic nanoarchitectures of different Morpho butterflies with different structural colors as biotemplates and examined the dependence of decomposition rates of methyl orange and rhodamine B dyes on the structural color of the biotemplates and the thickness of the ZnO coating. Using methyl orange, we measured a ten-fold increase in photodegradation rate when the 20 nm ZnO-coated wings were compared to similarly coated glass substrates. Using rhodamine B, a saturating relationship was found between the degradation rate and the thickness of the deposited ZnO on butterfly wings. We concluded that the enhancement of the catalytic efficiency can be attributed to the slow light effect due to a spectral overlap between the ZnO-coated Morpho butterfly wings reflectance with the absorption band of dyes, thus the photocatalytic performance could be changed by the tuning of the structural color of the butterfly biotemplates. The photodegradation mechanism of the dyes was investigated by liquid chromatography-mass spectroscopy.

Wide-gamut structural colours on oakblue butterflies by naturally tuned photonic nanoarchitectures.

The iridescent structural colours of butterflies, generated by photonic nanoarchitectures, often function as species-specific sexual signals; therefore, they are reproduced precisely from generation to generation. The wing scales of oakblue hairstreak butterflies (genus Arhopala, Theclinae, Lycaenidae, Lepidoptera) contain multi-layer photonic nanoarchitectures, which can generate a wide range of structural colours, from violet to green. By scanning (SEM) and cross-sectional transmission electron microscopy (TEM) investigation, the colour tuning mechanism of the cover scales was explored. We revealed that the characteristic size change of structural elements in similar photonic nanoarchitectures led to different structural colours that were examined by various reflectance spectrophotometry techniques. The measured structural properties of the naturally tuned photonic nanoarchitectures were used to calculate wing reflectances, which were compared with the measurement results. We found that the simulated structural colours were systematically redshifted by 95-126 nm as compared with the measured normal-incidence reflectance results. This is attributed to the swelling of the chitinous multi-layer structures during the standard TEM sample preparation and the tilt of the cover scales, which both affect the apparent layer thicknesses in the TEM cross-sections. We proposed a simulation correction and compared the results with the layer thicknesses measured on cryogenically prepared non-embedded SEM cross-sections.

Measuring and Modelling Structural Colours of Euphaedra neophron (Lepidoptera: Nymphalidae) Finely Tuned by Wing Scale Lower Lamina in Various Subspecies.

The nymphalid butterfly Euphaedra neophron (Hopffer, 1855) is the only structurally coloured species representing the genus along the Indian Ocean coast in East Africa and Southern Africa, with a distribution from southern Somalia to the Kwa-Zulu-Natal region of South Africa. The range of E. neophron is subdivided to several, geographically distinct populations, currently recognised as subspecies by taxonomists on the basis of violet, blue, and green-coloured morphs. We investigated the optical mechanism of all these morphs by various materials science techniques. We found that the structural colour is generated by the lower lamina of the cover scales and the different colours are tuned according to their thickness, which was also proved by modelling. The colour tuning of the different subspecies does not reflect any clinal pattern, be it geographical or altitudinal.

Spectral Engineering of Hybrid Biotemplated Photonic/Photocatalytic Nanoarchitectures.

Solar radiation is a cheap and abundant energy for water remediation, hydrogen generation by water splitting, and CO2 reduction. Supported photocatalysts have to be tuned to the pollutants to be eliminated. Spectral engineering may be a handy tool to increase the efficiency or the selectivity of these. Photonic nanoarchitectures of biological origin with hierarchical organization from nanometers to centimeters are candidates for such applications. We used the blue wing surface of laboratory-reared male Polyommatus icarus butterflies in combination with atomic layer deposition (ALD) of conformal ZnO coating and octahedral Cu2O nanoparticles (NP) to explore the possibilities of engineering the optical and catalytic properties of hybrid photonic nanoarchitectures. The samples were characterized by UV-Vis spectroscopy and optical and scanning electron microscopy. Their photocatalytic performance was benchmarked by comparing the initial decomposition rates of rhodamine B. Cu2O NPs alone or on the butterfly wings, covered by a 5 nm thick layer of ZnO, showed poor performance. Butterfly wings, or ZnO coated butterfly wings with 15 nm ALD layer showed a 3 to 3.5 times enhancement as compared to bare glass. The best performance of almost 4.3 times increase was obtained for the wings conformally coated with 15 nm ZnO, deposited with Cu2O NPs, followed by conformal coating with an additional 5 nm of ZnO by ALD. This enhanced efficiency is associated with slow light effects on the red edge of the reflectance maximum of the photonic nanoarchitectures and with enhanced carrier separation through the n-type ZnO and the p-type Cu2O heterojunction. Properly chosen biologic photonic nanoarchitectures in combination with carefully selected photocatalyst(s) can significantly increase the photodegradation of pollutants in water under visible light illumination.

Graphene-Encapsulated Silver Nanoparticles for Plasmonic Vapor Sensing.

Graphene-covered silver nanoparticles were prepared directly on highly oriented pyrolytic graphite substrates and characterized by atomic force microscopy. UV-Vis reflectance spectroscopy was used to measure the shift in the local surface plasmon resonance (LSPR) upon exposure to acetone, ethanol, 2-propanol, toluene, and water vapor. The optical responses were found to be substance-specific, as also demonstrated by principal component analysis. Point defects were introduced in the structure of the graphene overlayer by O2 plasma. The LSPR was affected by the plasma treatment, but it was completely recovered using subsequent annealing. It was found that the presence of defects increased the response for toluene and water while decreasing it for acetone.

Spectral tuning of biotemplated ZnO photonic nanoarchitectures for photocatalytic applications.

The photocatalytic activity of a flat surface can be increased by micro- and nanostructuring the interface to increase the area of the contact surface between the photocatalyst and the solute, and moreover, to optimize charge carrier transfer. Further enhancement can be achieved by using photonic nanostructures, which exhibit photonic band gap (PBG). Structurally coloured butterfly wings offer a rich 'library' of PBGs in the visible spectral range which can be used as naturally tuned sample sets for biotemplating. We used conformal atomic layer deposition of ZnO on the wings of various butterfly species (Arhopala asopia, Hypochrysops polycletus, Morpho sulkowskyi, Polyommatus icarus) possessing structural colour extending from the near UV to the blue wavelength range, to test the effects arising from the nanostructured surfaces and from the presence of different types of PBGs. Aqueous solutions of rhodamine B were used to test the enhancement of photocatalytic activity that was found for all ZnO-coated butterfly wings. The best reaction rate of decomposing rhodamine B when illuminated with visible light was found in 15 nm ZnO coated M. sulkowskyi wing, the reflectance of which had the highest overlap with the absorption band of the dye and had the highest reflectance intensity.

Large-area nanoengineering of graphene corrugations for visible-frequency graphene plasmons.

Quantum confinement of the charge carriers of graphene is an effective way to engineer its properties. This is commonly realized through physical edges that are associated with the deterioration of mobility and strong suppression of plasmon resonances. Here, we demonstrate a simple, large-area, edge-free nanostructuring technique, based on amplifying random nanoscale structural corrugations to a level where they efficiently confine charge carriers, without inducing significant inter-valley scattering. This soft confinement allows the low-loss lateral ultra-confinement of graphene plasmons, scaling up their resonance frequency from the native terahertz to the commercially relevant visible range. Visible graphene plasmons localized into nanocorrugations mediate much stronger light-matter interactions (Raman enhancement) than previously achieved with graphene, enabling the detection of specific molecules from femtomolar solutions or ambient air. Moreover, nanocorrugated graphene sheets also support propagating visible plasmon modes, as revealed by scanning near-field optical microscopy observation of their interference patterns.

Scale granules and colours: Sexual dimorphism in Trichonis (Lepidoptera: Lycaenidae, Theclinae).

A large fraction of dorsal wing surface ground scales show an unusual granulated nature, composed of material apparently extruded from the scale lumen in male individuals of both Trichonis Hewitson, 1865 species in the tribe Eumaeini, a rare Guyanian-Amazonian genus. Only a few not-granulated male specimens are known, females are not granulated. The granulated scales are investigated by various microscopic (optical, scanning and transmission electron microscopy, focused ion beam lamella cutting) and spectroscopic (optical reflectance, energy-dispersive X-ray (EDS), Raman) techniques. The characteristic blue colour unique in the South American representatives of the tribe is documented and analysed. EDS spectra show that the granules contain additional calcium and oxygen as compared with the un-granulated regions of the same scale. Electron diffraction (inside the TEM) did not reveal any crystalline component in the granules. The granulated wing surfaces of the males exhibit a UV absorption band at 280 nm, characteristic for biogenic CaCO3; therefore, the material of the granules is tentatively identified as CaCO3. It is shown that the granules influence the optical properties of the dorsal wing surface resulting in a characteristic spectrum.

Concordance of the spectral properties of dorsal wing scales with the phylogeographic structure of European male Polyommatus icarus butterflies.

The males of more than 80% of the Lycaenidae species belonging to the tribe Polyommatini exhibit structural coloration on their dorsal wing surfaces. These colors have a role in reinforcement in prezygotic reproductive isolation. The species-specific colors are produced by the cellular self-assembly of chitin/air nanocomposites. The spectral position of the reflectance maximum of such photonic nanoarchitectures depends on the nanoscale geometric dimensions of the elements building up the nanostructure. Previous work showed that the coloration of male Polyommatus icarus butterflies in the Western and Eastern Palearctic exhibits a characteristic spectral difference (20 nm). We investigated the coloration and the de novo developed DNA microsatellites of 80 P. icarus specimens from Europe from four sampling locations, spanning a distance of 1621 km. Remarkably good concordance was found between the spectral properties of the blue sexual signaling color (coincident within 5 nm) and the population genetic structure as revealed by 10 microsatellites for the P. icarus species.

Multi-instrumental techniques for evaluating butterfly structural colors: A case study on Polyommatus bellargus (Rottemburg, 1775) (Lepidoptera: Lycaenidae: Polyommatinae).

Color is an important communication channel for day-flying butterflies. Chemical (pigmentary) coloration is often supplemented by physical color generated by photonic nanostructures. These nanoarchitectures - which are characteristic for a given species - exhibit wavelength ranges in which light propagation is forbidden. The photonic nanoarchitectures are located in the lumen of the wing scales and are developed individually by each scale during metamorphosis. This self-assembly process is governed by the genes in the nucleus of the scale producing cell. It is crucial to establish well-defined measurement methods for the unambiguous characterization and comparison of colors generated in such a complex manner. Owing to the intricate architecture ordered at multiple levels (from centimeters to tens of nanometers), the precise quantitative determination of butterfly wing coloration is not trivial. In this paper, we present an overview of several optical spectroscopy measurement methods and illustrate techniques for processing the obtained data, using the species Polyommatus bellargus as a test case, the males of which exhibit a variation in their blue structural color that is easily recognizable to the naked eye. The benefits and drawbacks of these optical methods are discussed and compared. Furthermore, the origin of the color differences is explained in relation to differences in the wing scale nanomorphology revealed by electron microscopy. This in turn is tentatively associated with the unusually large genetic drift reported for this species in the literature.

Synthesis and Characterization of Graphene-Silver Nanoparticle Hybrid Materials.

Silver nanoparticles (Ag NPs) play important roles in the development of plasmonic applications. Combining these nanoparticles with graphene can yield hybrid materials with enhanced light-matter interaction. Here, we report a simple method for the synthesis of graphene-silver nanoparticle hybrids on highly oriented pyrolytic graphite (HOPG) substrates. We demonstrate by scanning tunneling microscopy and local tunneling spectroscopy measurements the electrostatic n-type doping of graphene by contact with silver. We show by UV-Vis reflectance investigations that the local surface plasmon resonance (LSPR) of Ag NPs partially covered with graphene is preserved for at least three months, i.e., three times longer than the LSPR of bare Ag NPs. The gradual loss of LSPR is due to the spontaneous sulfurization of non-covered Ag NPs, as revealed by scanning electron microscopy and energy-dispersive X-ray spectroscopy. We show that the Ag NPs completely sandwiched between graphene and HOPG do not sulfurize, even after one year.

Stability and Selective Vapor Sensing of Structurally Colored Lepidopteran Wings Under Humid Conditions.

Biological photonic nanoarchitectures are capable of rapidly and chemically selectively sensing volatile organic compounds due to changing color when exposed to such vapors. Here, stability and the vapor sensing properties of butterfly and moth wings were investigated by optical spectroscopy in the presence of water vapor. It was shown that repeated 30 s vapor exposures over 50 min did not change the resulting optical response signal in a time-dependent manner, and after 5-min exposures the sensor preserved its initial properties. Time-dependent response signals were shown to be species-specific, and by using five test substances they were also shown to be substance-specific. The latter was also evaluated using principal component analysis, which showed that the time-dependent optical responses can be used for real-time analysis of the vapors. It was demonstrated that the capability to detect volatile organic compounds was preserved in the presence of water vapor: high-intensity color change signals with short response times were measured in 25% relative humidity, similar to the one-component case; therefore, our results can contribute to the development of biological photonic nanoarchitecture-based vapor detectors for real-world applications, like living and working environments.

Reproducible phenotype alteration due to prolonged cooling of the pupae of Polyommatus icarus butterflies.

The phenotypic changes induced by prolonged cooling (2-12 weeks at 5 °C in the dark) of freshly formed Polyommatus icarus pupae were investigated. Cooling halted the imaginal development of pupae collected shortly after transformation from the larval stage. After cooling, the pupae were allowed to continue their developmental cycle. The wings of the eclosed specimens were investigated by optical microscopy, scanning and cross-sectional transmission electron microscopy, UV-VIS spectroscopy and microspectroscopy. The eclosed adults presented phenotypic alterations that reproduced results that we published previously for smaller groups of individuals remarkably well; these changes included i) a linear increase in the magnitude of quantified deviation from normal ventral wing patterns with increasing cooling time; ii) slight alteration of the blue coloration of males; and iii) an increasing number of blue scales on the dorsal wing surface of females with increasing cooling time. Several independent factors, including disordering of regular scale rows in males, the number of blue scales in females, eclosion probability and the probability of defect-free eclosion, showed that the cooling time can be divided into three periods: 0-4 weeks, 4-8 weeks, and 8-12 weeks, each of which is characterized by specific changes. The shift from brown female scales to first blue scales with a female-specific shape and then to blue scales with a male-specific shape with longer cooling times suggests slow decomposition of a substance governing scale formation.

High accuracy of color-generating nanoarchitectures is kept in lowland and mountainous populations of Polyommatus dorylas (Lepidoptera: Lycaenidae: Polyommatinae).

It is known that the size of the scales covering the surface of the Lepidoptera wings is in correlation with body size: larger species possess larger scales. However, butterfly individuals representing the various generations of the same species but differing in body size were not investigated in this respect. Similarly, the question whether different scale size may influence structural color generation based on nanoarchitectures in the scale lumen was never addressed. Populations of lowland (environment of Budapest, Hungary) and upland (Carpathian Mountains, Romania) Polyommatus dorylas were compared in terms of voltinism, wing and scale size, and the structural origin of blue coloration. Data analysis showed that the univoltine upland population exhibits a larger wing and scale size. On the other hand, the nanomorphology of the blue color-generating scales was identical when compared between univoltine and bivoltine populations. Coloration was also identical when measured with a spectrophotometer under ultraviolet and visible light. This high accuracy present in the male structural coloration suggests that it is controlled genetically. Body size alteration for enhanced thermal fitness has no influence on the fine structure of the nanoarchitecture present in the scale lumen.

Optical Detection of Vapor Mixtures Using Structurally Colored Butterfly and Moth Wings.

Photonic nanoarchitectures in the wing scales of butterflies and moths are capable of fast and chemically selective vapor sensing due to changing color when volatile vapors are introduced to the surrounding atmosphere. This process is based on the capillary condensation of the vapors, which results in the conformal change of the chitin-air nanoarchitectures and leads to a vapor-specific optical response. Here, we investigated the optical responses of the wing scales of several butterfly and moth species when mixtures of different volatile vapors were applied to the surrounding atmosphere. We found that the optical responses for the different vapor mixtures fell between the optical responses of the two pure solvents in all the investigated specimens. The detailed evaluation, using principal component analysis, showed that the butterfly-wing-based sensor material is capable of differentiating between vapor mixtures as the structural color response was found to be characteristic for each of them.

Modeling the Reflectance Changes Induced by Vapor Condensation in Lycaenid Butterfly Wing Scales Colored by Photonic Nanoarchitectures.

Gas/vapor sensors based on photonic band gap-type materials are attractive as they allow a quick optical readout. The photonic nanoarchitectures responsible for the coloration of the wing scales of many butterfly species possessing structural color exhibit chemical selectivity, i.e., give vapor-specific optical response signals. Modeling this complex physical-chemical process is very important to be able to exploit the possibilities of these photonic nanoarchitectures. We performed measurements of the ethanol vapor concentration-dependent reflectance spectra of the Albulina metallica butterfly, which exhibits structural color on both the dorsal (blue) and ventral (gold-green) wing sides. Using a numerical analysis of transmission electron microscopy (TEM) images, we revealed the details of the photonic nanoarchitecture inside the wing scales. On both sides, it is a 1D + 2D structure, a stack of layers, where the layers contain a quasi-ordered arrangement of air voids embedded in chitin. Next, we built a parametric simulation model that matched the measured spectra. The reflectance spectra were calculated by ab-initio methods by assuming variable amounts of vapor condensed to liquid in the air voids, as well as vapor concentration-dependent swelling of the chitin. From fitting the simulated results to the measured spectra, we found a similar swelling on both wing surfaces, but more liquid was found to concentrate in the smaller air voids for each vapor concentration value measured.

Biogeographical patterns in the structural blue of male Polyommatus icarus butterflies.

Color is a widely used communication channel in the living world for a variety of functions ranging from sexual communication to warning colors. A particularly rich spectrum of colors appears on the wings of many butterflies. The males of lycaenid butterflies often exhibit a conspicuous blue coloration generated by photonic nanoarchitectures on their dorsal wing surfaces. Using UV-VIS spectroscopy, we investigated the spatio-temporal variations of this coloration for Polyommatus icarus butterflies, considering an interval of more than 100 years and a geographical range spanning Europe (west) and Asia (east). The blue coloration in Hungary is very stable both within a year (three broods typical in Hungary) and within the period of 100 years (more than 300 generations). East-west geographic variation was investigated among 314 male P. icarus butterflies. In agreement with earlier genetic and morphometric studies, it was found that the western males are not divided in distinct lineages. Clear differences in coloration were found between the eastern and western groups, with a transition in the region of Turkey. These differences are tentatively attributed to bottleneck effects during past glaciations.

Vapour sensing properties of graphene-covered gold nanoparticles.

We investigated the vapour sensing properties of different graphene-gold hybrid nanostructures. We observed the shifts in the optical spectra near the local surface plasmon resonance of the gold nanoparticles by changing the concentration and nature of the analytes (ethanol, 2-propanol, and toluene). The smaller, dome-like gold nanoparticles proved to be more sensitive to these vapours compared to slightly larger, flat nanoparticles. We investigated how the optical response of the gold nanoparticles can be tuned with a corrugated graphene overlayer. We showed that the presence of graphene increased the sensitivity to ethanol and 2-propanol, while it decreased it towards toluene exposure (at concentrations ≥ 30%). The slope changes observed on the optical response curves were discussed in the framework of capillary condensation. These results can have potential impact on the development of new sensors based on graphene-gold hybrids.