A trade‐off between overheating and camouflage on shorebird eggshell colouration

In ground‐nesting birds egg colour and appearance may have evolved due to opposite selection pressures. Pigmentation and spottiness make the eggs darker and have been suggested to improve camouflage. However darker and more spotted eggs may reach higher temperatures when not attended by adults and receiving direct sunlight, which may be lethal for embryos. Some authors suggested that this trade‐off may not exist because eggshell pigments mainly reflect in the infrared region of the solar spectrum, but have not considered that wavelengths in the visible part of the spectrum may also contribute to overheating. To test the occurrence of a trade‐off between camouflage and overheating of eggs, we took digital images to analyse colour and camouflage in 93 nests of four shorebird species (two stilts and two plovers) in two regions (tropical and mediterranean sites). We predicted that these species (closely related) may have evolved different eggshell designs depending on solar radiation, which is supposed to be stronger in the Tropics. To record egg temperatures, we placed Japanese quail eggs in natural nests of shorebirds, and registered temperatures using a datalogger. We found that darker and more spotted eggs reached higher temperatures than lighter ones, and that after controlling for environmental temperatures, eggs overheated more in the Tropics, likely because of a more intense solar radiation. We also found that tropical shorebirds’ eggshells have darker spots and lighter backgrounds. Overall, darker eggs were better camouflaged. Taken together, our results show that the benefits of increasing pigmentation of eggshell backgrounds and spottiness for a better camouflage are counteracted by the increased risks of overheating when eggs remain exposed to direct solar radiation.     

Attractive blue‐green egg coloration and cuckoo−host coevolution

Recent evidence suggests that blue‐green coloration of bird eggshells may be related to female and/or egg phenotypic quality, and that such colour may affect parental effort and therefore the nutritional environment of developing nestlings. Here we suggest that these relationships and the signal function of eggshell coloration would affect the outcome of coevolution between avian brood parasites and their hosts in at least three different non‐exclusive evolutionary pathways. First, by laying blue‐green coloured eggs, cuckoo females may exploit possible sensory biases of their hosts, constraining the evolution of parasitic egg recognition, and thus avoid rejection. Second, because of the relatively high costs of laying blue eggs, cuckoo females may be limited in their ability to mimic costly blue‐green eggs of their hosts because cuckoo females lay many more eggs than their hosts. Furthermore, costs associated with foreign egg recognition errors would be relatively higher for hosts laying blue eggs. Third, cuckoos may use coloration of host eggs for selecting individuals or specific hosts of appropriate phenotypic quality (i.e. parental abilities). We here explored some predictions emerging from the above scenarios and found partial support for two of them by studying egg coloration of European cuckoos (Cuculus canorus) and that of their 25 main hosts, as well as parasitism and rejection rate of hosts. Cuckoo hosts parasitized with more blue, green, and ultraviolet cuckoo eggs, or those laying more blue‐green eggs, were more prone to accept experimental parasitism with artificial cuckoo eggs. In addition, coloration of cuckoo eggs is more variable when parasitizing hosts laying bluer‐greener eggs, even after controlling for the effect of host egg coloration (i.e. degree of egg matching). Globally, our results are consistent with the proposed hypothesis that host egg traits that are related to phenotypic quality of hosts, such as egg coloration, may have important implications for the coevolutionary interaction between hosts and brood parasites. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, 106 , 154–168.     

Dark nests and egg colour in birds: a possible functional role of ultraviolet reflectance in egg detectability

Owing to the conspicuousness of ultraviolet (UV) colour in dark environments, natural selection might have selected UV egg coloration because it would enhance egg detectability by parents in murky nests. Here, we tested this hypothesis by using comparative and experimental approaches. First, we studied variation in egg coloration of 98 species of European passerines measured using UV-visible reflectance spectrometry (300-700nm) in relation to nesting habits. Analyses based on raw data and controlling for phylogenetic distances both at the species and the family levels revealed that hole-nester species produced eggs with higher UV reflectance than those nesting in open habitats. The experimental approach consisted of the manipulation of UV reflectance of the experimental eggs introduced outside the nest-cup of the hole-nester spotless starling Sturnus unicolor and the study of the retrieval of these eggs. Ultraviolet-reflecting eggs (controls) were more frequently retrieved to the nest-cup than non-reflecting (-UV) eggs. These results were not due to '-UV' eggs being recognized by starlings as parasitic because when a parasitic egg is detected, starlings removed it from the nest-box. Therefore, these results are consistent with the hypothesis that UV egg colours are designed to provide highly detectable targets for parent birds in dark nest environments.     

Egg color as an adaptation for thermoregulation

ABSTRACT.   Avian embryos are incubated at temperatures only 2–6 °C below that at which hyperthermia begins to influence survival. In habitats where sunlight directly strikes the eggs, even for short periods, heat gain may be a substantial threat to survival, and reflective pigmentation may reduce the rate of heat gain. The results of previous studies suggest that light-colored eggs acquire heat slower than dark eggs, but artificial pigments were used to create differences in egg coloration. This approach is problematic because natural eggshell pigments have low absorbance in the near-infrared waveband that encompasses about half of incident solar radiation. We used naturally-pigmented eggs to measure the influence of egg coloration on heat gain. Triads ( N = 18) of eggs from Brewer's ( Euphagous cyanocephalus ), Red-winged ( Agelaius phoeniceus ), and Yellow-headed ( Xanthocephalus xanthocephalus ) blackbirds were crossed with six nests of each species and either exposed to full sunlight or placed under a diffusing umbrella. Thermisters recorded internal egg temperature every minute until an asymptotic temperature was reached. Eggs in full sunlight acquired heat more rapidly than eggs in the shaded environment, but heat gain did not vary with egg color in either environment. Eggs placed in Yellow-headed Blackbird nests took longer to reach asymptotic temperature, but there was no significant egg-by-nest interaction. Thus, it appears that differences in reflectivity of eggshell pigments in the visible range (400–700 nm) do not result in different rates of heat acquisition. The thermoregulation hypothesis was not supported.     

Incubation reduces microbial growth on eggshells and the opportunity for trans-shell infection

Avian eggshells harbour microbes shortly after laying, and under appropriate ambient conditions they can multiply rapidly, penetrate through shell pores, infect egg contents and cause embryo mortality. We experimentally examined how incubation affects bacterial processes on the eggshells of pearl-eyed thrashers Margarops fuscatus nesting in tropical montane and lowland forests in Puerto Rico. Bacteria and fungi grew rapidly on shells of newly laid, unincubated eggs exposed to ambient conditions, but declined to low levels on shells of eggs incubated by thrashers. Divergence in bacterial growth between incubated and exposed eggs was more marked at the montane forest than at the lowland site. Pathogenic microorganisms became increasingly dominant on shells of exposed eggs, but these groups were relatively rare on incubated eggs, where more benign, less invasive groups prevailed. Some incubation during laying may be necessary to decrease the probability of trans-shell infection by reducing the growth of harmful bacteria and fungi on eggshells, although it may increase hatching asynchrony and the likelihood of brood reduction.     

Egg Coloration and Recognition of Conspecific Brood Parasitism in Eastern Bluebirds

Individual eastern bluebird (Sialia sialis) females produce clutches of eggs with unique coloration and older females and females in better body condition lay more pigmented blue‐green eggs. Conspecific brood parasitism in this species is not uncommon and bluebirds occasionally reject what appear to be normal eggs by moving them to the periphery of the nest. I used UV‐visual reflectance spectrometry to objectively measure coloration of eggs and nest material. To estimate the conspicuousness of the trait, I calculated the contrast between eggs and background nest material. I found high achromatic and chromatic contrast between the coloration of eggs and of the nests, suggesting that bluebird eggs are highly conspicuous. To test the hypothesis that expression of blue‐green coloration eggs facilitates recognition of eggs laid by conspecific brood parasites, I cross‐fostered individual eggs into host nests during egg laying and monitored the fate of those eggs. I found no support, however, for the hypothesis that egg coloration facilitates discrimination of parasitic eggs from host eggs.     

Solar incubation cuts down parental care in a burrow nesting tropical shorebird,the crab plover Dromas ardeola

We describe an unknown mode of solar‐assisted egg development in the crab plover Dromas ardeola, a shorebird that breeds in self‐excavated burrows. The insulating properties of the nest burrow and the intense solar radiation allowed egg development at near‐optimal temperature (35.2°C±0.2) and humidity (60.2%±4.4), allowing a very low incubation attendance by the parent birds (28.3% of time, with recesses lasting up to 58 h). Crab plovers did not abandon completely parental incubation, possibly because of the need to turn their egg, and because the slight warming provided by parents (0.8°C) may improve hatching. This is the first case of solar assisted incubation in a species unrelated to the Megapodiidae, the only birds known to develop their eggs without contact incubation.     

Eggshells as hosts of bacterial communities: An experimental test of the antimicrobial egg coloration hypothesis

Oviparous animals have evolved multiple defenses to prevent microbes from penetrating their eggs and causing embryo mortality. In birds, egg constituents such as lysozyme and antibodies defend against microbial infestation, but eggshell pigments might also impact survival of bacteria. If so, microbes could exert an important selective pressure on the evolution of eggshell coloration. In a previous lab experiment, eggshell protoporphyrin caused drastic mortality in cultures of Gram positive, but not Gram negative, bacteria when exposed to light. Here, we test this “photodynamic antimicrobial hypothesis” in a field experiment. In a paired experimental design, we placed sanitized brown, protoporphyrin‐rich chicken eggs alongside white eggs that lack protoporphyrin. We deployed eggs for 48 hr without incubation, as can occur between laying and incubation, when microbial infection risk is highest. Eggs were placed on the open ground exposed to sunlight and in dark underground storm‐petrel burrows. We predicted that the proportion of Gram‐positive bacteria on brown eggs should be lower when exposed to sunlight than when kept in the dark, but we expected no such difference for white eggs. Although our data revealed variation in bacterial community composition, the proportion of Gram‐positive bacteria on eggshells did not vary by egg color, and there was no interaction between egg color and location. Instead, Gram‐positive bacteria were proportionally more common on eggs on the ground than eggs in burrows. Overall, our experiment did not support the photodynamic antimicrobial hypothesis. The diverse range of avian egg colors is generated by just two pigments, but over 10 hypotheses have been proposed for the evolution of eggshell color. If our results are generalizable, eggshell protoporphyrin might not play a substantial role in defending eggs against microbes, which narrows the field of candidate hypotheses for the evolution of avian eggshell coloration.     

Evidence of selection for egg crypsis in conspicuous nests

ABSTRACT The value of egg coloration as crypsis, once accepted as a general principle, has recently been questioned because most experiments have failed to show that egg coloration deters predation. The nest‐crypsis hypothesis postulates that, among species that build conspicuous nests, selection for egg crypsis is relaxed or absent because visually searching predators detect nests prior to eggs. I tested the nest‐crypsis hypothesis using the large, relatively conspicuous nests of American Robins (Turdus migratorius), and eggs that differed markedly in color that were collected from the nests of Red‐winged Blackbirds (Agelaius phoeniceus), Brewer's Blackbirds (Euphagus cyanocephalus), and Yellow‐headed Blackbirds (Xanthocephalus xanthocephalus). Each nest (N= 22) received a clutch of each species during three sequential predation trials that were 16 d in duration. The order of clutch presentation was randomized for each nest. Survival trends for Brewer's and Yellow‐headed Blackbirds were similar, and higher than those for clutches of Red‐winged Blackbirds. By the end of trials, overall survival of the three clutch types was roughly equivalent. However, clutches of Red‐winged Blackbird eggs, the most conspicuous egg type to the human eye, were discovered sooner by predators. Because the experimental design controlled for effects of nest crypsis, nest location, and nest size, this difference in egg survival can be attributed to differences in egg pigmentation. Thus, my results support a role for egg coloration as camouflage in conspicuous nests.     

The separate effects of egg size and parental quality on the development of ornamental plumage coloration

The fitness‐related consequences of egg size, independent of the influences of parental quality, are poorly understood in altricial birds. Not only can egg size and parental quality influence growth and survival, but each could influence the development of condition‐dependent plumage coloration in offspring. The Eastern Bluebird Sialia sialis is an altricial, multi‐brooded, cavity‐nesting passerine in which juveniles display dichromatic UV‐blue plumage. Previous research suggests that plumage coloration acts as a signal of individual quality among juvenile and adult Eastern Bluebirds. Here, we separate the effects of egg size and parental quality (defined by egg size laid) on nestling growth and plumage ornamentation by exchanging clutches of large eggs with clutches of small eggs. Nestlings were significantly larger immediately post‐hatching when hatched from a large egg, but to maintain a larger size, nestlings needed to have hatched from a large egg and to have been reared by high‐quality parents. Nestlings were brighter when reared by high‐quality parents and this relationship was strongest later in the breeding season. Nestlings exhibited greater UV chroma if hatched early in the season, but UV chroma was not significantly affected by egg size or parental quality. These findings demonstrate varying influences of both egg size and parental quality on offspring growth and plumage ornamentation but suggest that quality of post‐hatching investment is more influential than pre‐hatching investment.     

The functions of belly-soaking in Kentish Plovers Charadrius alexandrinus

We tested whether belly-soaking (i.e. wetting of ventral plumage) in incubating Kentish Plovers Charadrius alexandrinus in a hot environment serves to cool overheated eggs, or whether belly-soaking would mainly serve for adult thermoregulation, and egg wetting would be a mere consequence of the ventral feathers of incubating adults being wet. When ambient temperature was high (> 30 °C), body temperature of incubating Kentish Plovers increased with ambient temperatures. When adults departed from nests to belly-soak, egg temperatures were not critical for embryos. However, eggs in exposed nests overheated to levels that may be lethal for embryos when adults left the nests to belly-soak, and adults also employed belly-soaking to cool down overheated eggs when they returned to their nests. Indeed, the cooling rate of eggs was much faster when they were wetted. We conclude that the primary function of belly-soaking in the Kentish Plover is to dissipate body heat in heat-stressed incubating adults, rather than to cool overheated eggs. Eggs overheated as a consequence of short-term interruptions in nest attendance by heat-stressed adults to belly-soak, and Plovers employed secondarily belly-soaking to cool down overheated eggs. These results indicate that adult thermoregulation and egg cooling are not mutually exclusive functions of belly-soaking.     

Reflections on the origin of the amniote egg in the light of reproductive strategies and shell structure

On the basis of evolutionary pattern of reproductive strategies (r‐ and K‐selected animals) and the ultrastructure of modern amniote eggshells, a new model of the origin of the amniote egg is presented. In contrast to the well‐known idea of Romer (1957) that the egg came first while adult reptiles in the Carboniferous remained in water, it is suggested that the early evolution of the amniote egg (including the gradual formation of membranes) happened within the aquatic realm. Increasing enlargement of eggs and yolk is interpreted as an adaptation of reproduction strategies to lakes with poor nutrient contents. The first accumulation of Ca‐ions in the outer membrane, paralleled by many modern noncalcified lepidosaurian eggs, was a process of detoxification, according to new ideas in biomineralization. The function of the shell, to protect the embryo against microbial activity and to prevent water loss, which is necessary for the preamniote egg to become a fully terrestrial egg, was the terminal step in this story. Yolk‐rich eggs enclosed by a more or less calcified shell can be interpreted as an essential preadaptation for tetrapods to have become fully terrestrial during the Late Paleozoic.     

Egg shape and size allometry in geckos (Squamata: Gekkota), lizards with contrasting eggshell structure: why lay spherical eggs?

Hard, highly calcified eggshells evolved several independent times during the history of amniotes. Because of phylogenetic conservatism of this trait, lineages in which closely related taxa differ in eggshell structure are rare. Four gekkotan families (Carphodactylidae, Diplodactylidae, Eublepharidae and Pygopodidae) have eggs with soft shells, while their close relatives (Gekkonidae) lay eggs with hard shells. Geckos thus offer a rare opportunity to compare the impact of the emergence of a hard eggshell on the economy of egg architecture. Because a sphere has the smallest surface area of all three‐dimensional solids of a given volume, spherical eggs in geckos with hard eggshells reduce calcium investment and should therefore be advantageous. Here, we document that hard‐shelled gekkonid eggs are indeed more spherical than those of the other gecko lineages. However, within gekkonids, small species lay more elongated eggs than larger species. We speculate that miniature gekkonid females, which lay larger eggs relative to body size compared with large gekkonids, produce elongate eggs in order to pass the egg through a limited pelvic opening.     

Solar UV-B radiation affects below-ground parameters in a fen ecosystem in Tierra del Fuego, Argentina: implications of stratospheric ozone depletion

Stratospheric ozone depletion caused by the release of chlorofluorocarbons is most pronounced at high latitudes, especially in the Southern Hemisphere (including the so‐called ‘ozone hole’). The consequent increase in solar ultraviolet‐B radiation (UV‐B, 280–315 nm) reaching the earth's surface may cause a variety of alterations in terrestrial ecosystems. Most effects might be expected to occur above‐ground since sunlight does not penetrate effectively below‐ground. Here, we demonstrate that solar UV‐B radiation in a fen in Tierra del Fuego (Argentina), where the ozone hole passes overhead several times during the Austral spring, is causing large changes of below‐ground processes of this ecosystem. During the third and fourth year of a manipulative field experiment, we investigated root systems in these plots and found that when the ambient solar UV‐B radiation was substantially reduced, there was a 30% increase in summer root length production and as much as a threefold decrease in already low symbiotic mycorrhizal colonization frequency of the roots compared with plots receiving near‐ambient solar UV‐B. There was also an apparent shift toward older age classes of roots under reduced solar UV‐B. Such large changes in root system behaviour may have decided effects on competition and other ecological interactions in this ecosystem.     

产卵次序对山麻雀卵色、卵大小和重量的影响

产卵次序可对鸟类卵黄中的营养成分、卵壳色素以及卵壳厚度等产生影响。一些鸟类可通过产卵次序对不同卵的资源进行分配,从而实现繁殖成效的最大化。本研究对不同产卵次序下山麻雀(Passer cinnamomeus)的卵色、卵大小和卵重量进行分析,结果表明,山麻雀的卵重和卵大小随着产卵次序并无显著变化,卵色与卵的大小和重量均无显著相关性,但卵背景的色度随产卵次序有明显下降趋势,而卵斑点密度则呈相反的变化。卵背景的色度随产卵次序下降可能是山麻雀有限的色素在卵中不均匀分配的结果,而卵斑点密度的增加则可能是对后期卵壳厚度变薄的一种补偿。     

Spectral properties of plant leaves pertaining to urban landscape design of broad-spectrum solar ultraviolet radiation reduction

Human exposure to harmful ultraviolet (UV) radiation has important public health implications. Actual human exposure to solar UV radiation depends on ambient UV irradiance, and the latter is influenced by ground reflection. In urban areas with higher reflectivity, UV exposure occurs routinely. To discover the solar UV radiation regulation mechanism of vegetation, the spectral reflectance and transmittance of plant leaves were measured with a spectrophotometer. Typically, higher plants have low leaf reflectance (around 5%) and essentially zero transmittance throughout the UV region regardless of plant species and seasonal change. Accordingly, incident UV radiation decreases to 5% by being reflected and is reduced to zero by passing through a leaf. Therefore, stratified structures of vegetation are working as another terminator of UV rays, protecting whole terrestrial ecosystems, while vegetation at waterfronts contributes to protect aquatic ecosystems. It is possible to protect the human population from harmful UV radiation by urban landscape design of tree shade and the botanical environment. Even thin but uniformly distributed canopy is effective in attenuating UV radiation. To intercept diffuse radiation, UV screening by vertical structures such as hedges should be considered. Reflectivity of vegetation is around 2%, as foliage surfaces reduce incident UV radiation via reflection, while also eliminating it by transmittance. Accordingly, vegetation reduces incident UV radiation to around 2% by reflection. Vegetation influence on ambient UV radiation is broad-spectrum throughout the UV region. Only trees provide cool UV protective shade. Urban landscapes aimed at abating urban heat islands integrated with a reduction of human UV over-exposure would contribute to mitigation of climate change.     

Rethinking the thermal melanism hypothesis: rearing temperature and coloration in pygmy grasshoppers

Selection for efficient conversion of solar radiation to body heat has favored the evolution of dark coloration in many ectotherms. The thermal melanism hypothesis posits that dark coloration is beneficial under conditions of low ambient temperatures because it results in faster heating rates and higher body temperatures. Fast heating rates, however, may come at a cost of overheating unless compensated for by thermal physiology or behaviour. Pygmy grasshopper (Orthoptera, Tetrigidae) populations that inhabit fire-ravaged areas characterized by blackened backgrounds and hot surface temperatures due to high absorbance of solar radiation show an increased frequency of black phenotypes. I raised the progeny of wild-captured Tetrix undulata in cold and hot temperatures and used data on color patterns and survival in a greenhouse to examine whether a cold thermal environment triggered the development of melanic coloration or differently affected survival of melanic versus non-melanic individuals. My results indicate that melanism was not influenced by rearing temperature but by genes or epigenetic maternal effects. Temperature also did not affect survival. However, melanic individuals produced by melanic mothers survived longer than melanic individuals produced by non- melanic mothers, whereas non-melanic individuals produced by non-black mothers survived longer than melanic individuals produced by non-black mothers. This suggests a mismatch between color and physiology in offspring belonging to a different color morph than their mother. Future investigations into the evolution of melanism should consider conflicting selection pressures on thermal capacity and camouflage as well as the influence of correlated responses to selection on traits associated with coloration.     

The effect of embryonic development on metal and calcium content in eggs and eggshells in a small passerine

Published information relating to changes in the chemical element content of avian eggs caused by embryonic development is extremely scarce, although it may be crucial for understanding both the presence of anthropogenic pollutants as well as physiological levels of micronutrients. We assessed the variation in concentrations of calcium (Ca) and magnesium (Mg) and nine trace elements: seven essential (chromium (Cr), copper (Cu), nickel (Ni), manganese (Mn), iron (Fe), cobalt (Co) and zinc (Zn)) and two non‐essential (lead (Pb) and cadmium (Cd)) in shells and contents (both egg yolk and egg white) of embryonated and non‐embryonated eggs. We investigated the eggs of the Eurasian Reed Warbler Acrocephalus scirpaceus, a large proportion of whose eggs are infertile in our study population (almost 43% of clutches contain unhatched eggs) as well as significant embryo‐induced eggshell thinning at the equator of embryonated eggs. We found significantly higher concentrations (≥ 22.7%) of all the focal elements in the contents of embryonated eggs in comparison with non‐embryonated eggs, and a very pronounced one for Ca (nearly twice as high). The shells of embryonated eggs contained significantly higher concentrations of Zn (104.1%), Fe (56.5%), Pb (32.8%) and Cu (28.0%) but significantly lower ones of Co (8.9%) and Ca (9.3%) than the shells of non‐embryonated eggs. The simultaneous higher concentrations of all elements in the content of thinner‐shelled embryonated eggs suggest the parallel transfer of these elements along with Ca resorption from the shell into the egg interior during embryo formation. The higher concentration of most elements in the thinner shells of embryonated eggs may be indicative of the maternal deposition of some of these elements in a shell layer not subject to embryonic depletion, or in the eggshell membrane. Our results highlight the need for the careful selection of egg samples, which should differentiate between embryonated and non‐embryonated eggs in the analytical treatment of eggs and eggshells.     

Susceptibility of avian hosts to experimental Gymnophalloides seoi infection

To determine whether avian species are susceptible to infection with Gymnophalloides seoi (a human-infecting intestinal trematode), we exposed 7 species of birds with metacercariae obtained from oysters. The birds were necropsied at days 2, 4, and 6 postinfection (PI). The highest worm recovery at day 6 PI was obtained from the Kentish plover (Charadrius alexandrinus; mean = 56.0%), followed by the Mongolian plover (C. mongolus; 49.3%), and the grey plover (Pluvialis squatarola; 32.3%). In contrast, no mature worms were recovered from the great knot (Calidris tenuirostris), dunlin (C. alpina), black-tailed gull (Larus crassirostris), and mallard (Anas platyrhynchos). Among the plovers, the worms attained the greatest size at day 6 PI (254.1 x 190.4 microm) in the Kentish plover, with a significantly higher number of eggs in the uterus. The 3 species of plovers are highly susceptible to experimental G. seoi infection, suggesting that they could play a role as definitive hosts for these worms in nature.     

Eggshell Spottiness Reflects Maternally Transferred Antibodies in Blue Tits

Blue-green and brown-spotted eggshells in birds have been proposed as sexual signals of female physiological condition and egg quality, reflecting maternal investment in the egg. Testing this hypothesis requires linking eggshell coloration to egg content, which is lacking for brown protoporphyrin-based pigmentation. As protoporphyrins can induce oxidative stress, and a large amount in eggshells should indicate either high female and egg quality if it reflects the female''s high oxidative tolerance, or conversely poor quality if it reflects female physiological stress. Different studies supported either predictions but are difficult to compare given the methodological differences in eggshell-spottiness measurements. Using the blue tit Cyanistes caeruleus as a model species, we aimed at disentangling both predictions in testing if brown-spotted eggshell could reflect the quality of maternal investment in antibodies and carotenoids in the egg, and at improving between-study comparisons in correlating several common measurements of eggshell coloration (spectral and digital measures, spotted surface, pigmentation indices). We found that these color variables were weakly correlated highlighting the need for comparable quantitative measurements between studies and for multivariate regressions incorporating several eggshell-color characteristics. When evaluating the potential signaling function of brown-spotted eggshells, we thus searched for the brown eggshell-color variables that best predicted the maternal transfer of antibodies and carotenoids to egg yolks. We also tested the effects of several parental traits and breeding parameters potentially affecting this transfer. While eggshell coloration did not relate to yolk carotenoids, the eggs with larger and less evenly-distributed spots had higher antibody concentrations, suggesting that both the quantity and distribution of brown pigments reflected the transfer of maternal immune compounds in egg yolks. As yolk antibody concentrations were also positively related to key proxies of maternal quality (egg volume, number, yellow feather brightness, tarsus length), eggshells with larger spots concentrated at their broad pole may indicate higher-quality eggs.