The liver but not the skin is the site for conversion of a red carotenoid in a passerine bird

Carotenoids may provide numerous health benefits and are also responsible for the integumentary coloration of many bird species. Despite their importance, many aspects of their metabolism are still poorly known, and even basic issues such as the anatomical sites of conversion remain controversial. Recent studies suggest that the transformation of carotenoid pigments takes place directly in the follicles during feather growth, even though the liver has been previously recognised as a storing organ for these pigments with a certain potential for conversion. In this context, we analysed the carotenoid profile of plasma, liver, skin and feathers of male Common Crossbills (Loxia curvirostra). Interestingly, the derivative feather pigment 3-hydroxy-echinenone was detected in the liver and in the bloodstream (i.e. the necessary vehicle to transport metabolites to colourful peripheral tissues). Our results demonstrate for the first time with empirical data that the liver may act as the main site for the synthesis of integumentary carotenoids. This finding contradicts previous assumptions and raises the question of possible inter-specific differences in the site of carotenoid conversion in birds.     

Carotenoid-based colouration and ultraviolet reflectance of the sexual ornaments of grouse

Among the most familiar sexual signals are red, yellow, and orange sexual traits pigmented by carotenoids. Many birds can detect near-ultraviolet (UV) light, and UV signals can play key roles in mate choice. Grouse (Tetraonidae) exhibit bright carotenoid-dependent sexual ornaments, their supra-orbital combs, which to humans appear orange-red. Combs also reflect in the UV, which is not visible to humans but is likely to be visible to grouse. In male red grouse Lagopus lagopus scoticus, we show that comb UV reflectance decreases with increasing comb size and redness. By removing the epidermis of combs, where carotenoid pigments are, we show that the UV reflectance is a property of the dermis, underneath the red pigmented epidermis. Carotenoid pigmentation of combs acted as a mask to reduce reflectance by the dermis in the range 400–550 nm and in the UV, 300–400 nm. Patagium skin (non-ornamental skin under the wing) also reflects in the UV, but epidermis removal on this bare part tended to reduce UV reflectance, whereas removal of the red epidermis of combs increased UV reflectance. Males in better condition (greater body mass relative to size) had bigger and redder combs, but with less UV. Thus, carotenoid pigments of grouse combs are deposited on a white background with significant UV reflectance, which can influence how the signal is perceived by conspecifics. Carotenoid-based traits exhibit UV reflectance in a number of species, but how UV reflectance and carotenoid pigmentation influence colour remains little known for integumentary ornaments compared to plumage traits. UV vision is not uncommon in birds and other animals, so future studies should investigate how UV reflectance influences the perception of carotenoid-based signals of quality.     

Ornamental comb colour predicts T-cell-mediated immunity in male red grouse <Emphasis Type="Italic">Lagopus lagopus scoticus</Emphasis>

Sexual ornaments might reliably indicate the ability to cope with parasites and diseases, and a better ability to mount a primary inflammatory response to a novel challenge. Carotenoid-based ornaments are amongst the commonest sexual signals of birds and often influence mate choice. Because carotenoids are immuno-stimulants, signallers may trade-off allocating these to ornamental colouration or using them for immune responses, so carotenoid-based ornaments might be particularly useful as honest indicators of immuno-compentence. Tetraonid birds, such as the red grouse Lagopus lagopus scoticus, exhibit supra-orbital yellow-red combs, a conspicuous ornament which functions in intra- and inter-sexual selection. The colour of combs is due to epidermal pigmentation by carotenoids, while their size is testosterone-dependent. In this study, I investigated whether comb characteristics, and in particular, comb colour, indicated immuno-competence in free-living male red grouse. I assessed T-cell-mediated immunity using a standardised challenge with phytohaemagglutinin. Red grouse combs reflect in the red and in the ultraviolet spectrum of light, which is not visible to humans but that grouse most likely see, so I measured comb colour across the whole bird visible spectrum (300-700 nm) using a reflectance spectrometer. I found that males with bigger and redder combs, but with less ultraviolet reflectance, had greater T-cell-mediated immune response. Comb colour predicted T-cell-mediated immune response better than comb size, indicating that the carotenoid-based colouration of this ornament might reliably signal this aspect of male quality.     

Testosterone upregulates lipoprotein status to control sexual attractiveness in a colorful songbird

A salient feature of many secondary sexual characteristics in animals is that their expression is controlled by sex-steroid hormones. However, for only a few types of ornaments do we know the precise molecular mechanism by which androgens like testosterone (T) enhance trait production. We studied the red carotenoid-based beak of male zebra finches (Taeniopygia guttata), which serves as a sexually selected trait and is thought to be T dependent. In previous research, we demonstrated that the extent to which male finches produce lipoproteins in the bloodstream, which bind carotenoid pigments acquired from the diet and transport them to peripheral tissues, regulates the accumulation of carotenoids in the body and beak pigmentation. Here we show that T acts to upregulate lipoprotein production and allows male zebra finches to display flashy, sexually attractive coloration. Levels of circulating T in blood positively and significantly predicted lipoprotein profile (as measured by cholesterol levels), blood carotenoid concentration, and beak color. Exogenous T administration elevated cholesterol and carotenoid status as well as beak redness. Last, experimental inhibition of T (using the anti-androgen flutamide) downregulated lipoprotein production and carotenoid circulation and faded the beak. This androgen- and lipoprotein-mediated system represents one of the more detailed physiological mechanisms underlying the development of a sex-steroid-dependent trait in animals.     

Enhancement of Rhodobacter sphaeroides growth and carotenoid production through biostimulation

Bacillus thuringiensis/cereus L2 was added as a biostimulant to enhance the biomass accumulation and carotenoid yield of Rhodobacter sphaeroides using wastewater as the culturing medium. Results showed that biostimulation could significantly enhance the R. sphaeroides biomass production and carotenoid yield. The optimal biostimulant proportion was 40 μL(about 6.4 × 105CFU). Through the use of biostimulation, chemical oxygen demand removal, R. sphaeroides biomass production, carotenoid concentration, and carotenoid yield were improved by 178%, 67%, 214%, and 70%, respectively. Theoretical analysis revealed that there were two possible reasons for such increases. One was that biostimulation enhanced the R. sphaeroides wastewater treatment efficiency. The other was that biostimulation significantly decreased the peroxidase activity in R. sphaeroides. The results showed that the highest peroxidase activity dropped by 87% and the induction ratio of the RSP_3419 gene was 3.1 with the addition of biostimulant. The enhanced carotenoid yield in R. sphaeroides could thus be explained by a decrease in peroxidase activity.     

A quantitative comparison of the commonly used methods for extracting carotenoids from avian plasma

Interest in animal carotenoids, especially in birds, has exploded in recent years, and so too have the methods employed to investigate the nature and function of these pigments. Perhaps the most easily and commonly performed procedure in this work has been the determination of carotenoid concentration from avian plasma. Over the past 20 years of research on avian carotenoids, numerous methods have been used to extract carotenoids from bird plasma, all of which have differed in several important parameters (e.g., number and types of solvents used, degree of mixing/centrifugation). However, to date, no study has systematically compared these methods to determine if any of them are more effective than others for recovering any or all types of carotenoids present. We undertook such an investigation on plasma samples from two bird species (house finch, Carpodacus mexicanus, and mallard, Anas platyrhynchos) using five of the most commonly employed methods for extracting carotenoids from avian plasma: (1) acetone-only, (2) methanol-only, (3) ethanol-only, (4) ethanol + hexane, and (5) ethanol + tert butyl methyl ether. We also manipulated the amount of time that extracts were centrifuged, which has varied tremendously in previous studies, to evaluate its importance on carotenoid recovery. We found that all methods equally recovered the polar xanthophylls (lutein and zeaxanthin), but that the methanol-only procedure poorly recovered non-polar carotenoids (less β-carotene in both species and less β-cryptoxanthin in house finches) compared to the other methods. These results suggest that the data accumulated to date on xanthophyll plasma carotenoids in birds should be comparable across studies and species despite the different chemical extraction methods used. However, care should be taken to use relatively strong organic solvents for fully recovering non-polar carotenoids. We also found no effect of centrifugation duration (1 vs. 10 min at 10,000 rpm) on carotenoid recoveries, demonstrating that researchers can save considerable time by centrifuging for a much shorter time period than is typically used.     

Tolerance of an Albino Fish to Ultraviolet-B Radiation

We exposed albino and pigmented medakaOryzias latipes to simulated solar ultraviolet-B (UVB) radiation to determine if albino medaka were less tolerant of UVB radiation than medaka pigmented with melanin. There was no difference in the number of albino and pigmented medaka that died during the exposure period. Spectrophotometric analyses of the outer dorsal skin layers from albino and pigmented medaka indicated that, prior to exposure, both groups of fish had similar amounts of an apparent colorless non-melanin photoprotective substance that appears to protect other fish species from UVB radiation. Our results indicate that albino medaka were as tolerant of UVB radiation as pigmented medaka because they had similar amounts of this photoprotective substance in the outer layers of the skin.     

Immunoregulatory activity of different dietary carotenoids in male zebra finches

Summary. Many animals use carotenoid pigments to color their integument and become sexually attractive. These colorants can also serve physiological functions, protecting cells and tissues from oxidative damage as well as stimulating the immune system. Because animals often acquire several different carotenoid pigments from their diet, there is the potential for different carotenoids to exhibit different free-radical-scavenging or immunoenhancing activity. We experimentally tested how two common dietary xanthophylls - lutein and zeaxanthin - may differentially affect the immune system in male zebra finches (Taeniopygia guttata). Male T. guttata derive their red sexual beak colorants from these two carotenoids, and prior studies with this species have shown that lutein and zeaxanthin together boost cell-mediated immunity. We experimentally elevated these two dietary carotenoids separately in two groups of non-breeding zebra finches, but found that lutein-supplemented and zeaxanthin-supplemented males mounted similar cell-mediated immune responses (to phytohaemagglutinin, or PHA). Although zeaxanthin is a more conjugated carotenoid than lutein and has the potential to be a more potent antioxidant, our study suggests that such a subtle structural difference between these two biochemicals does not differentially affect immune performance in this songbird.     

Egg coloration is correlated with female condition in eastern bluebirds (Sialia sialis)

Egg coloration has been hypothesized to reflect female condition. Because of the proposed physiological costs associated with deposition of biliverdin pigments and because of their conspicuousness, eggs with blue-green coloration may reliably convey information about female or brood quality. We tested the hypothesis that expression of blue-green coloration of eastern bluebird (Sialia sialis) eggs positively correlates to female condition. First, we documented the incidence of egg color polymorphism within the population. We observed that 98% of females laid blue-green eggs while less than 2% laid white eggs and less than 1% laid pink eggs. In a subset of clutches, we used full spectrum reflectance spectrometry (300–700 nm) to compare eggshell coloration to measures of female condition. We found that the color of eggs within clutches was more similar than the color of eggs from different clutches, and that the blue-green eggs have spectral peaks that are consistent with the characteristic absorbance spectra of biliverdin pigmentation. Females in better body condition and older females laid more colorful eggs. Moreover, individual females laid more colorful eggs later in the laying sequence. Overall, these data indicate that egg coloration covaries with female condition, suggesting that egg coloration could function as a reliable signal of female quality or that egg coloration may allow females to recognize eggs laid by conspecific brood parasites.     

Carotenoid-derived ornaments reflect parental quality in male and female yellow-eyed penguins (<Emphasis Type="Italic">Megadyptes antipodes</Emphasis>)

The handicap principle suggests that ornamental traits that function as honest signals in mate selection must be costly to be effective. We evaluated in the sexually monochromatic yellow-eyed penguin (Megadyptes antipodes) whether the carotenoid-derived plumage and eye coloration predicts parental quality and whether males and females within pairs mate assortatively in relation to these carotenoid-derived ornaments. In addition, we investigated whether age or body condition was related to the coloration of the ornamental traits. In yellow-eyed penguins, parental quality of males and females was predicted by eye and head plumage coloration. Even when we controlled for gender- and age-specific differences, eye and head plumage coloration reflected honestly parental quality. Males and females mated assortatively in relation to these ornamental traits. While age influenced coloration of both the eye and head plumage, body condition was related only to the saturation of plumage coloration. These results provide evidence that the carotenoid-derived ornaments in yellow-eyed penguins reflect the parental abilities of birds and, therefore, may be costly signals. Potentially, female and male yellow-eyed penguins could use eye and plumage coloration as an indirect cue in assessing age and quality of individual birds during mate choice. This is only the second study to examine plumage coloration in relation to sexual selection in penguins, while conspicuous ornamental traits in other species of penguin beg the question whether they also play a role in sexual selection.Electronic Supplementary Material Supplementary material is available in the online version of this article at Communicated by C.R. Brown