Summary1. Recent technical and methodological advances have led to a dramatic increase in the use of spectrometry to quantify reflectance properties of biological materials, as well as models to determine how these colours are perceived by animals, providing important insights into ecological and evolutionary aspects of animal visual communication. 2. Despite this growing interest, a unified cross-platform framework for analysing and visualizing spectral data has not been available. We introduce pavo, an R package that facilitates the organization, visualization and analysis of spectral data in a cohesive framework. pavo is highly flexible, allowing users to (a) organize and manipulate data from a variety of sources, (b) visualize data using R's state-of-the-art graphics capabilities and (c) analyse data using spectral curve shape properties and visual system modelling for a broad range of taxa. 3. In this paper, we present a summary of the functions implemented in pavo and how they integrate in a workflow to explore and analyse spectral data. We also present an exact solution for the calculation of colour volume overlap in colourspace, thus expanding previously published methodologies. 4. As an example of pavo's capabilities, we compare the colour patterns of three African glossy starling species, two of which have diverged very recently. We demonstrate how both colour vision models and direct spectral measurement analysis can be used to describe colour attributes and differences between these species. Different approaches to visual models and several plotting capabilities exemplify the package's versatility and streamlined workflow. 5. pavo provides a cohesive environment for handling spectral data and addressing complex sensory ecology questions, while integrating with R's modular core for a broader and comprehensive analytical framework, automated management of spectral data and reproducible workflows for colour analysis.

The colours of birds are diverse but limited relative to the colours they can perceive. This mismatch may be partially caused by the properties of their colour-production mechanisms. Aside from pigments, several classes of highly ordered nanostructures (thin films, amorphous three-dimensional arrays) can produce a range of colours. However, the variability of any single nanostructural class has rarely been explored. Dabbling ducks are a speciose clade with substantial interspecific variation in the iridescent coloration of their wing patches (specula). Here, we use electron microscopy, spectrophotometry, polarization and refractive index-matching experiments, and optical modelling to examine these colours. We show that, in all species examined, speculum colour is produced by a photonic heterostructure consisting of both a single thin-film of keratin and a two-dimensional hexagonal lattice of melanosomes in feather barbules. Although the range of possible variations of this heterostructure is theoretically broad, only relatively close-packed, energetically stable variants producing more saturated colours were observed, suggesting that ducks are either physically constrained to these configurations or are under selection for the colours that they produce. These data thus reveal a previously undescribed biophotonic structure and suggest that both physical variability and constraints within single nanostructural classes may help explain the broader patterns of colour across Aves.

The Jurassic Yanliao theropods have offered rare glimpses of the early paravian evolution and particularly of bird origins, but, with the exception of the bizarre scansoriopterygids, they have shown similar skeletal and integumentary morphologies. Here we report a distinctive new Yanliao theropod species bearing prominent lacrimal crests, bony ornaments previously known from more basal theropods. It shows longer arm and leg feathers than Anchiornis and tail feathers with asymmetrical vanes forming a tail surface area even larger than that in Archaeopteryx. Nanostructures, interpreted as melanosomes, are morphologically similar to organized, platelet-shaped organelles that produce bright iridescent colours in extant birds. The new species indicates the presence of bony ornaments, feather colour and flight-related features consistent with proposed rapid character evolution and significant diversity in signalling and locomotor strategies near bird origins.

Developmental constraints and trade-offs can limit diversity, but organisms have repeatedly evolved morphological innovations that overcome these limits by expanding the range and functionality of traits. Iridescent colours in birds are commonly produced by melanin-containing organelles (melanosomes) organized into nanostructured arrays within feather barbules. Variation in array type (e.g. multilayers and photonic crystals, PCs) is known to have remarkable effects on plumage colour, but the optical consequences of variation in melanosome shape remain poorly understood. Here, we used a combination of spectrophotometric, experimental and theoretical methods to test how melanosome hollowness-a morphological innovation largely restricted to birds-affects feather colour. Optical analyses of hexagonal close-packed arrays of hollow melanosomes in two species, wild turkeys (Meleagris gallopavo) and violet-backed starlings (Cinnyricinclus leucogaster), indicated that they function as two-dimensional PCs. Incorporation of a larger dataset and optical modelling showed that, compared with solid melanosomes, hollow melanosomes allow birds to produce distinct colours with the same energetically favourable, close-packed configurations. These data suggest that a morphological novelty has, at least in part, allowed birds to achieve their vast morphological and colour diversity.

Evolutionary conflict in trait performance under different ecological contexts is common, but may also arise from functional coupling between traits operating within the same context. Orb webs first intercept and then retain insects long enough to be attacked by spiders. Improving either function increases prey capture and they are largely determined by different aspects of web architecture. We manipulated the mesh width of orbs to investigate its effect, along with web size, on prey capture by spiders and found that they functioned independently. Probability of prey capture increased with web size but was not affected by mesh width. Conversely, spiders on narrow-meshed webs were almost three times more likely to capture energetically profitable large insects, which demand greater prey retention. Yet, the two functions are still constrained during web spinning because increasing mesh width maximizes web size and hence interception, while retention is improved by decreasing mesh width because more silk adheres to insects. The architectural coupling between prey interception and retention has probably played a key role in both the macroevolution of orb web shape and the expression of plasticity in the spinning behaviours of spiders.

Infection is an important source of mortality for avian embryos but parental behaviors and eggs themselves can provide a network of antimicrobial defenses. Mound builders (Aves: Megapodiidae) are unique among birds in that they produce heat for developing embryos not by sitting on eggs but by burying them in carefully tended mounds of soil and microbially decomposing vegetation. The low infection rate of eggs of one species in particular, the Australian brush-turkey (Alectura lathami), suggests that they possess strong defensive mechanisms. To identify some of these mechanisms, we first quantified antimicrobial albumen proteins and characterized eggshell structure, finding that albumen was not unusually antimicrobial, but that eggshell cuticle was composed of nanometer-sized calcite spheres. Experimental tests revealed that these modified eggshells were significantly more hydrophobic and better at preventing bacterial attachment and penetration into the egg contents than chicken eggs. Our results suggest that these mechanisms may contribute to the antimicrobial defense system of these eggs, and may provide inspiration for new biomimetic anti-fouling surfaces.

Understanding how animal signals are produced is critical for understanding their evolution because complexity and modularity in the underlying morphology can affect evolutionary patterns. Hummingbird feathers show some of the brightest and most iridescent colors in nature. These are produced by optically complex stacks of hollow, platelet‐shaped organelles called melanosomes. Neither how these morphologies produce colors nor their evolution has been systematically studied. We first used nanoscale morphological measurements and optical modeling to identify the physical basis of color production in 34 hummingbird species. We found that, in general, the melanosome stacks function as multilayer reflectors, with platelet thickness and air space size explaining variation in hue (color) and saturation (color purity). Additionally, light rays reflected from the outer keratin surface interact with those reflected by small, superficial melanosomes to cause secondary reflectance peaks, primarily in short (blue) wavelengths. We then compared variation of both the morphological components and the colors they produce. The outer keratin cortex evolves independently and is more variable than other morphological traits, possibly due to functional constraints on melanosome packing. Intriguingly, shorter wavelength colors evolve faster than longer wavelength colors, perhaps due to developmental processes that enables greater lability of the shapes of small melanosomes. Together, these data indicate that increased structural complexity of feather tissues is associated with greater variation in morphology and iridescent coloration.

The avian eggshell is a highly ordered structure with several layers (mammillae, palisades, and vertical crystal layer) composed of calcium carbonate (∼96%) and minerals within an organic matrix. The cuticle is a noncalcified layer that covers the eggshells of most bird species. Eggshells are multifunctional structures that have evolved in response to diverse embryonic requirements and challenges, including protection from microbial infection, nest flooding, and exposure to solar radiation. However, experimental evidence for these functions across diverse taxa is currently limited. Here we investigated the effects of nanosphere cuticles on (1) bacterial attachment and transshell penetration, (2) eggshell wettability, (3) water vapor conductance, and (4) regulation of ultraviolet (UV) reflectance in seven ground-nesting bird species. We found considerable interspecific variation in ultrastructure and chemical composition of cuticles. Experimental removal of the cuticle confirmed that all nanospheres were highly effective at decreasing attachment of bacteria to shell surfaces and at preventing bacterial penetration. Cuticles also greatly decreased the amount of UV reflected by eggshells. In species with particularly small nanospheres, gas exchange was reduced by the presence of cuticle. Our results support the hypothesis that microbes and solar UV radiation can cause strong selection on bird eggs but also show that we need a greater understanding about the effects of specific nesting conditions (e.g., hydric and gaseous milieu) on embryo well-being and eggshell structure variation.