INTRODUCTION

Carotenoid pigments are responsible for many of the brightest colours and most conspicuous signals in birds (Hill 1999, Hill 2002). However, birds are not capable of synthesizing carotenoids from basic biological precursors (Goodwin 1984), and therefore, the expression of carotenoid pigments as colour signals requires the ingestion of carotenoids with food, as well as its absorption, transport, sometimes metabolism and incorporation into the feathers, all processes that are generally considered costly to individuals (Hill 2002, McGraw et al. 2005, McGraw et al. 2006). Scarcity of carotenoids in the food, or even the energetic cost of enzyme production and manipulation of carotenoids, remain unclear. Carotenoids are also involved in metabolic pathways related to host immunity and a trade-off between the ornamental and health functions of carotenoids has been proposed (Hõrak et al. 2004a, Møller et al. 2004). According to this hypothesis, signals based on carotenoid pigments are costly and therefore act as honest indicators of an individual's quality.

The features that each carotenoid-based ornament reflect have been mainly studied in males of very dimorphic species. Carotenoid-based ornaments have been shown to be related to body condition (McGraw et al. 2002, Saks et al. 2003, Jawor & Breitwisch 2004, Jawor et al. 2004), sexual selection (Collias et al. 1979, Hill 1991, Drachman 1997) and even parasite loads (Figuerola et al. 2003, Hill et al. 2004, Hõrak et al. 2004). In the case of parasites, studies have reported variously negative (Thompson et al. 1997, Brawner et al. 2000, Figuerola et al. 2003, Hõrak et al. 2004b), positive (Burley et al. 1991, VanHoort 8: Dawson 2005) and non-significant (Seutin 1994) relationships between parasites and the expression of a carotenoid-based ornament. The potential causes of these contradictory results could stem from varying methods of study (e.g. correlational vs. experimental, ranges or doses of experimental treatments used), different impacts on parasites on their hosts and environmental-dependent effects of parasites on host condition (Figuerola et al. 2003). Work published up to now has generally focused on a single type of parasite, above all blood parasites, and analysis of the relative and combined impact of different types of parasites on ornament expression is still lacking.

Haematozoan parasites are blood-cell parasite protozoa transmitted by blood-sucking arthropods that are quite prevalent in wild passerines (Deviche et al. 2001). Coccidian protozoa are widespread intestinal epithelium parasites with a direct biological cycle; transmission results from the ingestion of oocysts liberated in the faeces of an infected individual. Passerines are mainly infected by genus Isospora (Hill 2002, Hõrak et al. 2006). Both haematozoan and coccidian parasites affect their hosts in a condition-dependent way (Merino et al. 2000): they have little impact when resources are abundant (Weatherhead & Bennett 1992, Friend & Franson 2001), but affect negatively the host when resources are scarce (Ots & Hõrak 1998, Ilmonen et al. 1999). The mechanisms leading Coccidia to limit the expression of carotenoid-based ornaments may work in at least two different direct ways. First, they may reduce the absorption of carotenoids through the intestine (Tyczkowski et al. 1991, Allen 1992) and, second, they may reduce the release of high-density lipoproteins (Allen 1987), which are responsible for the transport of carotenoids in the bloodstream to the tissues. Moreover, a third indirect mechanism related to body condition – to which both carotenoid ornamentation and immunity have shown to be linked – may also be at work (see Smith et al. 2007). The mechanisms through which Haematozoa may limit the expression of carotenoid-based ornaments are still unknown.

Most of the relationships described up to now between parasites and the expression of ornaments have been made focusing on conspicuous male ornaments, but little attention have been traditionally paid to such a relation in female ornaments. Roulin (2001), for instance, conducted an experimental study by comparing the degree of female Barn Owl Tyto alba ornamentation (eumelanin-based spottiness) with parasite resistance in their offspring raised by foster parents. He found that in females ornamentation positively reflects parasite resistance ability. Some observational studies have also demonstrated that the expression of ornaments is negatively related to the parasite load in female birds (Hõrak et al. 2001, Piersma et al. 2001).

The aim of this study was to explore the relationship between the colour of the carotenoid-based red mask of European Goldfinches, and a number of indices of condition (haematological parameters and different parasite loads) in both sexes, paying attention to possible inter-sex differences. The study was carried out with free-living birds during the breeding season, just after mate choice had occurred, when individuals were going through the costly task of raising young, because under these conditions we expected the effect of parasites on their hosts to be maximal. To our knowledge, this is the first study done analysing the relationship between plumage coloration in both sexes and several groups of parasites at a time.

METHODS

The European Goldfinch is a 12-cm long, seed-eating finch that has a unique colour pattern on its head. The front of the face has a conspicuous crimson patch, which is known to be composed of four carotenoid pigments (Stradi et al. 1995): a) ε,ε-carotene-3,3′-dione, b) 3-hydroxy-ε,ε-carotene-3′-one, c) 4,4′-dihydroxy-ε,ε-carotene-3,3′-dione (isoastaxanthin), and d) 4-hydroxy-ε,ε-carotene-3,3′-dione.

Although ε,ε-carotene-3,3′-dione and 3-hydroxy-ε,ε-carotene-3′-one are very common yellow pigments in cardueline finches, isoastaxanthin and 4-hydroxy-ε,ε-carotene-3,3′-dione have not yet been found in any other species studied to date. These two pigments provide the red colour in the mask together with the keratin bond arrangement the pigments have in the feathers (Stradi et al. 1995). Although both sexes are superficially similar (Cramp 8: Perrins 1994), small differences in the size of the patch exist, being larger on average in males (Svensson 1996). To our knowledge, differences between the sexes in mask colour have never been investigated.

Colour characteristics

Reflectance curves were analysed by a Principal Component Analysis of reflectances at the 39 different wavelength intervals. Four relevant components (Eigenvalues > 1) were obtained that together summarised 99.97% of variance (Fig. 1). The first component (PC1) summarised reflection in the visible spectrum, mainly between 400 and 530 nanometers (within violets and blues). PC2 was more positive for individuals with more reflection at 650–740 nanometers (reds) and less reflection at lower wave lengths. PC3 was negatively related to reflection in the 550–590 nanometers intervals (yellows), so this component represents the yellow carotenoids reflection. PC4 was mainly influenced by reflection in the non-visible-to-humans portion of the spectra, between 360 and 390 nanometers (UVA radiation). The repeatability of the colour measurements was calculated as the intra-class correlation of the principal components from ten individuals measured twice (Lessells & Boag 1987). The repeatability was very high for all components (PC1: 98%; PC2: 96%; PC3: 97%; PC4: 96%) because of the great accuracy of the spectrometer method (see Figuerola et al. 1999).

Figure 1.

Factor loading of the four principal components calculated from light reflectance at 10 nm intervals between 360 and 740 nm. PC1 represents the blue-violet component, PC2 the red one, PC3 includes the yellows, and PC4 represents the UVA reflection.

RESULTS

The coloration of the red mask in the European Goldfinch was sexually dichromatic. Sexes differed in reflectance along the whole visible spectrum (PC1, PC2 and PC3), especially within the yellow region (PC3), but not in the UV (PC4) (Table 1).

Table 1.

Mean, SE and sample size for males and females for the colour, parasites and haematological variables analysed. Differences between sexes were tested by one-way ANOVA.

Reflectance curves showed that males were on average much more red and less yellow than females (Fig. 2). No differences in haematological values or parasite infection were found between sexes (Table 1). None of these variables were related to PC1 or PC2 (Tables 2 and 3). Haemoproteus infection, TLC, and their interaction with sex were related to PC3 (Tables 2 and 3, Fig. 3A and B). Isospora infection, its interaction with sex, and the interaction between sex and Haemoproteus infection were related to PC4 (Tables 2 and 3, Fig. 3C).

Figure 2.

Mean reflection curves of the red mask of the Goldfinch along the UV and the whole visible spectrum by sex.

Table 2.

Results of stepwise backwards selection procedure MANOVA analysing parasite infection over colour components of the red mask of the Eurasian Goldfinch.

DISCUSSION

Colour dichromatism has not been reported before in the masks of the European Goldfinch. Our results show that hues differ between sexes: males reflect reds more strongly than females, but reflect yellows and oranges with lower intensity than females. The carotenoids expressed in the mask, are qualitatively the same in both sexes (Stradi 1995). McGraw et al. (2002) also found that male American Goldfinches Carduelis tritis artificiallyfed with ad libitum canthaxantin were more colourful than females, due to a higher carotenoid concentration in the feathers. The larger accumulation of red pigments in males than in females seems thus to be the most plausible option for explaining colour differences in European Goldfinches. Testosterone, by means of its capacity to upregulate lipoprotein status, has been proposed as the responsible agent for such differences in the American Goldfinch (McGraw et al. 2006), but there are also studies showing opposite outcomes in House Finches Carpodacus mexicanus (Stoehr & Hill 2001). Diet differences between sexes, health variations or the effect of other hormones should not be discarded to explain this sexual dichromatism. Even a differential selection of ornaments between sexes could also be an underlying factor regarding the dichromatism. Unfortunately, no information is available on any of these aspects in the Eurasian goldfinch.

Table 3.

Results of stepwise backwards selection procedure MANOVA analysing haematological values over colour components of the red mask of the Eurasian Goldfinch.

Figure 3.

A) Scatter graph of log-transformed TLC over the PC3 values, by sex. B) Mean PC3 in relation to Haemoproteus infection state by sex. C) Mean PC4 in relation to Isospora infection state by sex.

When interpreting our results relating coloration to health it is important to consider that the study was carried out in the spring. European Goldfinches moult their masks in late summer (Jenni & Winkler 1994), around six months before reproduction takes place. The health status of individuals is expected to change during this time, although the signals involved in sexual selection act in the early spring at the time of mate choice (Cramp et al. 1994). The red of the mask is not fully developed at the time of the moult and is only completed during the spring due to the abrasion of melanin derived feather tips (Svensson 1996). We sampled birds at the time of the year when the expression of the ornament is at its fullest, just when the indicator function of a sexual selection signal should be at work.

We found a relationship between presence of different parasites and different colour components of red masks. However, these effects were sex-dependent and only significant for females. Female European Goldfinches infected with Haemoproteus blood parasites and those with higher TLC values were more orange, that is with a higher yellow component, than those uninfected or with lower values. A higher intensity in yellow component may be due to 1) a decrease in the intensity of red pigments, or 2) an increase in the intensity of yellow ones. Because red pigments are predominant in the red mask, we think that the first option is more plausible than the second one. In this way, the red of the mask may reflect Haemoproteus parasitemia or infection resistance in females. Also, female European Goldfinches with higher TLC values were more orange (with a higher yellow component) than those with lower values. Since high TLC values are linked to chronic or acute infections (Campbell 1995), the red of the mask may reflect immune levels or infection resistance in females. In this way, the most infected females would have less red ornaments, a finding that suggests that red masks act as an honest indicator of general infection in female European Goldfinches. This relationship is not significant in males, probably due to the different reproductive role of each sex, since egg-laying and incubation, a very expensive process, is carried out only by females (Cramp et al. 1994). UV reflection was higher in Isospora non-infected females than in infected ones in our study. This result seems to indicate that UV reflection acts as an honest indicator of Isospora infection in females. Finally, our results suggest that double-infected animals (with Haemoproteus and Isospora) reflect violets with a higher intensity that those non-infected. This fact could be due to the lack of red and yellow pigments (carotenoids) that those individuals have in their feathers. Although H/L has shown to be related with some aspects of stress and condition in passerines (Ots et al. 1998, Groombridge et al. 2004), it was not related to any colour variables in our study.

How is it possible that breeding roles had an effect on the relationship between coloration and parasites if plumage was developed several months before breeding? We suggest that under the stress derived from breeding activities females in worst condition or with a less active immune system are less able to control already present infections and/or exclude new infections when exposed to pathogens. A similar process was experimentally demonstrated to work in male Green-finches Carduelis chloris experimentally infected with Sindbis virus (Lindström & Lundstrom 2000).

In conclusion, our study shows that (1) sexual dichromatism exists in the colour of the mask of the European Goldfinch, and (2) the red colour of the mask reflects different signals in both sexes and may be a reliable indicator of parasite infection during the breeding season, at least in females.