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Colour characteristics of the blunt egg pole: cues for recognition of parasitic eggs as revealed by reflectance spectrophotometry
ANIMAL BEHAVIOUR, 2007, 74, 419e427 doi:10.1016/j.anbehav.2006.10.023
Colour characteristics of the blunt egg pole: cues for recognition of parasitic eggs as revealed by reflectance spectrophotometry OVA ´ *†, M ARC EL H ONZ A* , PETR P ROCH A ´ ZK A* , JA ´ N TOP ERCER‡ & B A˚ R D G. ST OKK E§ LENKA POLA CIK
*Institute of Vertebrate Biology, v.v.i, Academy of Sciences of the Czech Republic yInstitute of Botany and Zoology, Masaryk University in Brno zBotanical Garden, Comenius University xDepartment of Biology, Norwegian University of Science and Technology (Received 12 May 2006; initial acceptance 28 June 2006; final acceptance 16 October 2006; published online 22 August 2007; MS. number: 8958R)
Variation in appearance of parasitic and host eggs is an essential trait in the coevolutionary struggle between a brood parasite and its host. Here we examined the role of colour characteristics in the egg recognition in blackcaps, Sylvia atricapilla, assessed by (1) human vision and (2) spectrophotometry, including UV wavelengths. Since colour and spotting pattern within eggs are often highly variable, we also assessed three different parts of blackcap eggshell separately. We investigated the possibility that a lower intraclutch variation in the parasitized clutch would positively affect egg recognition. In addition, we predicted that egg rejection would depend on contrast between host and parasitic eggs. However, we found that neither intraclutch variation nor mimicry if assessed by humans had significant effect on host rejection. Multivariate analyses of spectrophotometric data revealed, however, that the likelihood of egg rejection significantly increased with decreasing intraclutch variation in blue chroma. Furthermore, probability of egg rejection significantly increased with decreasing contrast between parasitic and host eggs in UV wavelengths and with decreasing brightness of host eggs at the blunt egg pole. Accepted foreign eggs were significantly darker at the blunt pole than acceptors’ own eggs. The paradox that rejected eggs had lower contrast could be explained by the fact that the contrast in UV spectra is associated with brightness of the blunt egg part. Our data suggest that colour characteristics of the blunt egg part may play a major role in the recognition of parasitic eggs in the blackcaps. Ó 2007 The Association for the Study of Animal Behaviour. Published by Elsevier Ltd. All rights reserved.
Keywords: blackcap; brood parasitism; recognition cues; reflectance spectrophotometry; rejection behaviour; Sylvia atricapilla
Various aspects of bird life histories affect the colour and spotting pattern of eggshells (Kilner 2006). Coloration of the bird eggshell varies between and within species, ranging from achromatic pale to diversely spotted eggs (Makatsch 1976; Gosler et al. 2000). Recently, Underwood & Sealy (2002) and Kilner (2006) reviewed several Correspondence: L. Polacikova´, Institute of Vertebrate Biology, v.v.i, Academy of Sciences of the Czech Republic, Kveˇtna´ 8, CZ-603 65 Brno, Czech Republic and the Institute of Botany and Zoology, Masaryk University in Brno, Kotla´ˇrska´ 2, CZ-611 19 Brno, Czech Republic (email: [email protected]). J. Topercer is at the Botanical Garden, Comenius University, SK-038 15 Blatnica, Slovakia. B. G. Stokke is at the Department of Biology, Norwegian University of Science and Technology, NTNU, NO-7491 Trondheim, Norway. 0003e 3472/07/$30.00/0
hypotheses explaining the significance of variability in the colours and patterns on bird eggs. For example, egg coloration may allow clutches to mimic the environment and thus reduce the risk of nest predation (Solı´s & de Lope 1995). The egg colour pattern may also facilitate birds identifying their own clutches in colonially nesting birds (Birkhead 1978). Another notable functional explanation of avian egg coloration is related to the evolution of defences against brood parasitism, allowing hosts to recognise and eject foreign eggs in own clutches. On the other hand, brood parasite may evolve eggs that mimic those of their hosts in both ground colour and spotting pattern (Davies 2000). Brood parasitism in birds provides one of the best models for studying coevolutionary processes consisting
419 Ó 2007 The Association for the Study of Animal Behaviour. Published by Elsevier Ltd. All rights reserved.
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ANIMAL BEHAVIOUR, 74, 3
of the reciprocal defences and counterdefences which lead to a coevolutionary ‘arms race’ between hosts and brood parasites (Dawkins & Krebs 1979; Davies & de Brooke 1989). An interspecific brood parasitic reproductive strategy is deployed by about 1% of all bird species (Payne 1997), and comprises laying eggs in nests of another species, the hosts, which incubate the eggs and feed the parasitic young (Lack 1968). If the parasitic act is successful, it generally has detrimental effect on the hosts’ fitness because their reproductive success will be close to zero. This phenomenon represents a strong evolutionary selective force for the hosts to evolve counterdefences enabling them to overcome the negative effects of the parasite (Davies 2000). Several host adaptations related to egg appearance facilitate recognition of the parasitic egg. It has been shown that lower intraclutch and higher interclutch variation in appearance of host eggs (Øien et al. 1995; Stokke et al. 1999, 2002) and especially the contrast between host and parasitic eggs (Davies & de Brooke 1989; Moksnes et al. 1990; Honza et al. 2004; Stokke et al. 2004; Antonov et al. 2006) are the most relevant predictors for acceptance or rejection of foreign eggs. Some previous studies investigated the host ability to assess variation within own clutches based on human perception directly (Stokke et al. 1999; Welbergen et al. 2001; Procha´zka & Honza 2004) or based on image analysis software in the human visible spectrum (Soler et al. 2000). However, several studies assessed egg similarity by reflectance spectrophotometry, because there are anatomical differences between avian and human eyes (Bowmaker et al. 1997; Vorobyev et al. 1998; Cuthill et al. 1999). Birds differ from humans in colour perception (Bennett & Cuthill 1994), their cones being sensitive to a wider spectral range, which enables them perception of wavelengths below 400 nm (Goldsmith 1991). Indeed, recent studies have shown that birds use their ability to perceive ultraviolet wavelengths in mate choice (Bennett et al. 1997; Cuthill et al. 1999) and foraging (Honkavaara et al. 2002). Spectrophotometry techniques have also been applied in studies of avian brood parasitism, and several authors have pointed out the role of UV reflectance in egg recognition (Cherry & Bennett 2001; Soler et al. 2003; Underwood 2003; Avile´s & Møller 2004). However, Avile´s et al. (2006) found that artificial reduction of reflectance in the UV region of great spotted cuckoo, Clamator glandarius, eggs did not affect the ejection of parasitic eggs by their magpie, Pica pica, hosts. In this study, we tested the effect of colour characteristics on parasitic egg rejection in the blackcap, Sylvia atricapilla. This species is a good candidate for such kind of study because its eggs are apparently (based on human vision) highly variable in ground colour and marking pattern between clutches with differently coloured spots, speckles and blotches, which occasionally concentrate on the blunt pole (Makatsch 1976). Furthermore, there is evidence that blackcaps have been used as hosts by the common cuckoo, Cuculus canorus, in the past (Berthold et al. 1995; Moksnes & Røskaft 1995; Honza et al. 2001). As a consequence of this, blackcaps have retained good recognition ability (Moksnes & Røskaft 1992; Honza et al. 2004), and
generally have a low intraclutch and high interclutch variation in egg appearance (Øien et al. 1995). Honza et al. (2004) in a previous study showed that intraclutch variation did not influence rejection responses of the blackcaps. In the light of the above facts, we put forward two predictions. Since this species shows low intraclutch variation, we hypothesized that the intraclutch variation in this species would not significantly affect egg rejection in the hosts as found by Honza et al. (2004). Moreover, we also predicted that egg rejection would depend on contrast between host and parasitic eggs. To test the predictions, we studied the effect of variation in egg appearance within host clutches and degree of mimicry of the introduced egg on rejection behaviour by a traditional method based on human vision and by an objective spectrophotometry analysis. It has already been documented by traditional subjective methods that blackcap rejection behaviour is affected by specific colour characteristics (Honza et al. 2004). However, more objective spectrophotometric techniques can reveal in more detail which part of the light spectrum is important for rejection of foreign eggs. Furthermore, even though the avian egg has a complex colour pattern, previous spectrophotometric studies used mean values of reflectance per egg as a whole. To overcome this, we analysed both mean values per egg and three separate parts of the eggshell (sharp, medium, and blunt part) to test whether certain regions of the egg possess important cues for egg recognition that could be hidden if reflectance from all parts of the egg was averaged. METHODS
Experimental Procedure The study was carried out in a deciduous forest near Dolnı´ Bojanovice (48 510 N, 17 020 E), Czech Republic from 30 April to 22 May 2005. Blackcaps and cuckoos live in sympatry in this area, but we did not record any case of interspecific parasitism during our study. We searched systematically for nests during laying period or the first days of incubation. Eggs found during incubation were floated to estimate their approximate laying date according to methods described by Hays & LeCroy (1971). The eggs were measured (length, L; breadth, B) to the nearest 0.1 mm using callipers. Egg volume (V) was calculated using the formula V ¼ 0.51 L B2 (Hoyt 1979). All eggs were marked by small numbers on the medium part of the eggshell using waterproof ink to allow individual identification. No host egg was rejected, therefore is unlikely that the numbers had any significant effect on egg recognition by the hosts. Blackcap clutches were artificially parasitized either by adding a conspecific egg to the clutch without manipulating the original number of host eggs or exchanging one host egg with a conspecific egg. Altogether 44 experiments were carried out. Conspecific eggs are ideal for their use in this type of research because they are highly variable, ranging from good to poor mimicry. In addition, conspecific eggs are more suitable than model eggs because they are real (models are often deserted), variable in coloration, have natural range of coloration including
ET AL.: EGG RECOGNITION IN BLACKCAPS POLA CIKOV A
in UV, which would be extremely difficult if impossible to mimic using artificial colours. In total, 22 eggs were added and 22 eggs were exchanged during laying period only when at least the fourth host egg was laid. The nests were monitored daily for 5 days after manipulation. If the parasitic egg disappeared within this period but the host eggs were left unharmed, it was considered to have been rejected. If the parasitic egg remained unharmed in the nest, it was considered to have been accepted. If the clutch was abandoned within the 5-day observation period, it was considered to have been deserted. Three depredated clutches were excluded from the analyses.
Estimation of Intraclutch Variation and Contrast from Photographs The whole clutch including the parasitic egg was photographed on a grey background card with KODAK colour scale using a Nikon F65 camera and Fujicolor 200 ASA film. The intraclutch variation in egg appearance and the contrast between parasitic and host eggs were later assessed from the photos by 10 test persons, who judged both whole eggs and each egg part (blunt, medium and sharp) separately (for estimate of contrast), and were unaware of the fate of the introduced eggs. The intraclutch variation in egg ground colour and spotting pattern was scored on the following scale from one to five (Øien et al. 1995). (1) No variation, all the eggs were similar. (2) At least one egg differed slightly from the others. (3) At least one egg showed marked differences from the other eggs. (4) At least one egg differed dramatically from the others. (5) No two eggs were similar. The contrast between the parasitic and host eggs was scored on the following scale from one to three (Braa et al. 1992). (1) No contrast between host and parasitic eggs. The parasitic egg was indistinguishable from the host eggs. (2) Medium contrast between host and parasitic eggs. The parasitic egg could be distinguished from the host eggs, but the difference was only moderate. (3) High contrast between host and parasitic eggs. The parasitic egg could easily be distinguished from the host eggs. The median of scores by the test persons who assessed the whole eggs and the egg parts was used as estimate of intraclutch variation and contrast for each clutch. There was significant repeatability in scoring of the whole eggs among the 10 observers in intraclutch variability (Kendall coefficient of concordance: W43 ¼ 0.55, P < 0.0001) and in contrast between host and foreign eggs (W43 ¼ 0.56, P < 0.0001). Similarly, there was significant repeatability in scoring the contrast between host and parasitic eggs among the 10 observers for the blunt egg parts (W43 ¼ 0.69, P < 0.0001), the medium egg parts (W43 ¼ 0.60, P < 0.000) and the sharp egg parts (W43 ¼ 0.61, P < 0.0001).
Measurement of the Reflectance Spectra We measured the spectral reflectance of all the eggs from the parasitized clutches in the range 300e700 nm using a USB2000 spectrophotometer (Ocean Optics). We divided
each egg into three regions across the longitudinal axis of the egg. Each region comprised a third of the length of the egg representing the blunt, medium and sharp parts of the egg. From each egg part three randomly located measurements were taken (each covering 1 mm2). The illuminant was a deuterium and halogen light source (DT-Mini-GS, Ocean Optics). The light was transferred to the eggs through a quartz optic fibre (QR400-7-UV/VIS-BX, Ocean Optics), reached the eggs and was reflected from the eggs at an angle of 45 to the surface. Data from the spectrophotometer were loaded into OOIBase 32 software (Ocean Optics). The measurements were relative and referred to a standard white reference (WS-2) and to darkness. Reference and dark calibration were made before measurement of each clutch. Total reflectance was obtained for the UV (325e400 nm) interval, which comprises the human invisible part of colour spectrum, and for the blue (400e475 nm), green (475e550 nm), yellow (550e625 nm) and red (625e700 nm) wavelengths as the human visible regions. We calculated objective variables of the colour, namely brightness (reflectance intensity), chroma (purity of a colour) and hue (peak wavelengths) (Endler 1990). Brightness was calculated as the sum of the total reflectance values within the 325e 700 nm range of wavelengths (R325e700). Chroma was calculated as reflectance ratios: R325e400/325e700 (UV chroma), R400e475/325e700 (blue chroma), R475e550/325e700 (green chroma), R550e625/325e700 (yellow chroma) and R625e700/325e700 (red chroma). Hue was estimated by the wavelength of the maximum reflectance (Rmax). We performed the measurements on 193 eggs from 44 clutches. Despite large variation in spottiness of eggs we found significant repeatability for all nine reflectance measurements of each egg (Pearson correlation coefficients: for all 193 eggs: r > 0.80, P < 0.0001). Similarly, we found significant repeatability for all colour variables calculated for each egg (Sokal & Rohlf 1997; for all colour variables P < 0.001, df ¼ 192,1736; Table 1). Therefore we used mean values for each part of the colour spectrum per egg.
Estimation of Intraclutch Variation and Contrast from Spectrophotometric Measurements To estimate the degree of intraclutch variation we calculated coefficient of variation (SD/mean 100) from Table 1. Estimated repeatability (Pearson correlation coefficients) for colour characteristics of nine spectrophotometric measurements per each blackcap egg Colour characteristic Brightness Chroma UV Chroma blue Chroma green Chroma yellow Chroma red Hue
Repeatability
F192,1736
0.24*** 0.32*** 0.37*** 0.28*** 0.58*** 0.37*** 0.53***
3.77 5.25 6.00 4.45 13.5 6.2 11.21
193 Eggs from 44 clutches, ***P < 0.001.
421
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mean values for the whole egg of all colour variables in each clutch (Avile´s & Møller 2003). To estimate the contrast in coloration between host and parasitic eggs we calculated absolute differences of mean reflectance value in the ultraviolet, blue, green, yellow and red wavelengths among all hosts and parasitic eggs in the same clutch. Similarly, we calculated the absolute differences of mean reflectance values for blunt, medium and sharp part of all hosts and parasitic eggs in the same clutch separately. Mean values of these absolute differences were considered to represent the contrast between host and parasitic eggs. Finally, we compared mean total intensity of light (brightness) reflected from all eggs in each clutch as well as mean reflected light intensity for each egg part of all eggs in each clutch including brightness of accepted and rejected parasitic eggs.
Statistical Analyses To assess differences between the two experimental groups in response to parasitic eggs we used Fisher’s exact test, and for detecting differences in the laying day between acceptors and rejecters we used ManneWhitney U test. Most of the values acquired by scoring the photos (60%) significantly departed from normal distribution (D’AgostinoePearson omnibus test: P < 0.05). Because of this fact and the multivariate nature of the problem, we took advantage of multiple logistic regression with logit link (Hintze 1997e2004; Sokal & Rohlf 1997) to explore simultaneously the effects of intraclutch variation and contrast between host and parasitic eggs and their three parts on rejection behaviour. As all scores were recorded on the ordinal scale of measurement and the arithmetic mean is applicable only to ratio or interval scale data (Zar 1996), we submitted medians instead of means to the analysis. The comparison of reflected light intensity of all eggs in each clutch fully met the assumption of normality (D’AgostinoePearson omnibus test: P > 0.05), whereas the mean values of colour variables of intraclutch variation obtained by spectrophotometry and variables related to differences between parasitic and host eggs in the same nest violated the normality assumption in ca. 71% and 90% of cases, respectively (P < 0.05). Again we used multiple logistic regression to analyse the effects of intraclutch variation and differences in the contrast between host and foreign eggs on rejection behaviour and to assess the relative importance of these effects. To make the effect magnitudes comparable, we converted the values of all colour variables to the same scale of size and variability (from 0 to 1) using transformation proposed by Gower (Legendre & Legendre 1983). We used nonparametric tests for comparing brightness of eggs among egg parts separately (ManneWhitney U test, KruskaleWallis ANOVA test and Bonferroni post hoc test). All analyses proceeded using the statistical systems NCSS 2004 (Hintze 1997e2004) and Statistica 6.0 (StatSoft Inc. 1984e2001, Tulsa, OK, U.S.A.).
Ethical Note Because the eggs were measured under standard light conditions in the laboratory, we exchanged temporarily the host clutch for four to five foreign blackcap eggs to prevent nest desertion. In all nests the birds resumed incubation and none of these eggs was ejected. After the arrival to the lab, eggs were placed in an incubator (Octagon 20, Brinsea, U.K.), with temperature ranging from 37.7 to 37.8 C and relative humidity from 55% to 65%. The eggs were returned to the original nest within 1 h. Twenty-six (59.1%) nests were controlled until hatching. Of these, 14 were predated and 12 survived until hatching and in these nests all eggs hatched. Thus, we are sure that our temporal egg removal had no detrimental effect on hatching success. Moreover, survival to hatching of experimental nests was even slightly higher (Mayfield method, daily survival rate 0.96) compared to a control group of 41 nests (daily survival rate 0.91). All eggs used for experiments were freshly laid and rejection occurred soon after egg addition, that is, in the first days of incubation (incubation period in the blackcap is 12 days). Experiments were conducted in accordance with current laws of the Czech Republic and the Academy of Sciences Animal Care Protocol, licence number: 0008/98-M103.
RESULTS
Experimental Groups In the total of 44 nests, 28 (63.6%) introduced eggs were accepted and 16 (36.4%) eggs were rejected. There were no significant differences in response to alien eggs between the experimental groups in which the parasitic egg was added or exchanged with one of the host eggs (Fisher’s exact test: N1 ¼ 22, N2 ¼ 22, P ¼ 0.75). Also, there was no significant difference in the time to rejection of the parasitic egg between the two experimental groups (Manne Whitney U test: U ¼ 23, N1 ¼ 7, N2 ¼ 9, P ¼ 0.24, median 1 day (interquartile range: IQR 0e2 days), median 1 day (IQR 0e3 days)). Finally, there was no significant difference between the average size of host and the parasitic eggs added to the clutch (mean SD ¼ 2163.51 235.97 mm3, N1 ¼ 101 versus 2454.98 1665.68 mm3, N2 ¼ 22; U ¼ 1017, P ¼ 0.54) or exchanged for one host egg (2106.45 227.61 mm3, N1 ¼ 71 versus 2072.24 244.25 mm3, N2 ¼ 22; U ¼ 672, P ¼ 0.32). Therefore, we pooled the two experimental groups in further analyses.
Intraclutch Variation and Contrast between Host and Parasitic Eggs Assessed by Humans Of the five variables entered into the initial logistic regression model (medians of intraclutch variation and contrast in whole egg and medians of contrasts in three egg parts), none indicated significant effect on reaction (reaction ¼ 2, i.e. rejection) (c21 ¼ 0.37, P ¼ 0.544). Even the best model, containing medians of contrast in whole egg and in the middle part as predictor variables, was not significant (deviance ¼ 56.49, c22 ¼ 2.22, P ¼ 0.33).
ET AL.: EGG RECOGNITION IN BLACKCAPS POLA CIKOV A
Intraclutch Variation and Contrast between Host and Parasitic Eggs Based on Spectrophotometry Of the 31 candidate colour variables, we omitted 14 due to multicollinearity. The rest entered the hierarchical forward subset selection of variables up to one-way interactions with switching (Hintze 1997e2004). We used it in a stepwise manner, that is, we began with the best-fit model with one variable (best single predictor) then built the best-fit model with two variables and so on. Seven logistic regression models were fitted with 13 variables in total (see Table 2), while remaining four variables did not contribute significantly to any model (contrast in coloration of medium egg part at UV and yellow wavelengths, contrast in sharp egg part at blue wavelengths and average brightness; deviance tests: P > 0.2). Each model proved to be highly significant and valid except for that with six variables, in which maximum likelihood algorithm failed to converge and which was thus replaced by the second model. Except for contrast in blunt part at red wavelengths and average contrast at blue wavelengths, each variable showed significant effect on egg rejection behaviour in at least one model (deviance tests: P < 0.05). Contrast in coloration of blunt part at UV wavelengths (also the best single predictor: c21 ¼ 12.35, P ¼ 0.0004), intraclutch variation in blue chroma and brightness of blunt part were the most useful variables in prediction, gaining significance in six, five and four valid models, respectively. Their regression coefficients had consistent negative values similar in size ratios across the models, indicating (1) inverse relationships to rejection behaviour (the likelihood of alien egg rejection increases with decreasing values of both contrast in blunt part at UV wavelengths, intraclutch variation in blue chroma and brightness of blunt part) and (2) roughly equal effect magnitudes of the three variables. A trend towards a positive correlation between two of them (contrast in blunt part at UV wavelengths versus brightness of blunt part: Spearman rank correlation: rS ¼ 0.26, N ¼ 44, P ¼ 0.084) arose from additional correlation analysis. The simplest model for estimating the probability of egg rejection in blackcap using the three most productive predictor variables is: p2 ¼
e½5:464:91ðCh Blue ivÞ4:54ðUV epc1Þ5:16ðBrightness epb1Þ ; 1 þ e½5:464:91ðCh Blue ivÞ4:54ðUV epc1Þ5:16ðBrightness epb1Þ
where p2 is the probability of an egg being rejected, e is the base of natural logarithms, Ch_Blue_iv is the intraclutch variation in blue chroma, UV_epc1 is the contrast in coloration of blunt part at UV wavelengths and Brightness_ epb1 is the brightness of blunt part. Model accuracy was 77.27%, suggesting quite good ability to predict the egg rejection behaviour. Its explanatory power, however, was not that high (R2 ¼ 0.36). To explain the negative regression coefficients relating the rejection behaviour to the absolute differences between host and parasitic eggs at the blunt egg part, we looked closer at the brightness of the introduced and host eggs. The multiple logistic regression based on
spectrophotometry showed that the brightness reflected from the blunt egg part was lower in rejecters than acceptors (Fig. 1), a result consistent with two-sample testing (t42 ¼ 2.13, N1 ¼ 28, N2 ¼ 16, P ¼ 0.02; mean SD ¼ 24 174.58 4195.60 versus 27 465.11 5290.10). When we compared brightness of the blunt parts of accepted and rejected parasitic eggs, we did not confirm any significant differences although accepted eggs tended to be darker than rejected eggs (ManneWhitney U test: U ¼ 157, N1 ¼ 28, N2 ¼ 16, P > 0.05; mean SD ¼ 24 253.83 7992.30 versus 26 245.49 4358.75). Third, we compared the brightness of rejected eggs with the host eggs and we found that blunt egg parts of host clutches were darker than rejected eggs although the difference was not significant (U ¼ 98, N1 ¼ 16, N2 ¼ 16, P > 0.05; 24 174.58 4195.61 versus 26 245.49 4358.75). Finally, when we compared the brightness of accepted eggs with the host eggs, we found that blunt egg parts of host clutches were significantly lighter than accepted eggs (U ¼ 252, N1 ¼ 28, N2 ¼ 28, P ¼ 0.02; 27 465.64 5289.61 versus 24 253.83 7992.30). To confirm the differences between individual parts of an egg, we additionally compared also the colour variables of blunt, medium and sharp parts of all eggs. When using Bonferroni test, we found significant differences between all three egg parts in all colour characteristics (for all P < 0.016, N ¼ 579), except that blunt and medium egg parts did not differ significantly in yellow chroma (P ¼ 0.99).
DISCUSSION The blackcap shows exquisite egg recognition ability and rejects nearly 100% of nonmimetic eggs from its nests (Honza et al. 2004). The majority of individuals are therefore apparently able to reject foreign eggs as long as the appearance of foreign eggs is above a specific threshold for egg recognition based on individual’s cognitive abilities (Stokke et al. 2005). In line with our prediction, logistic regression model analysing the photo scores did not reveal significant influence of intraclutch variation on the host response. Similarly, many previous studies found the same pattern (Procha´zka & Honza 2003, 2004; Honza et al. 2004; Stokke et al. 2004). However, interestingly, our models based on spectrometric measurements revealed that the probability of egg rejection is negatively associated with intraclutch variation in host egg appearance in one part of the spectrum. Birds that rejected parasitic eggs showed lower variation in blue chroma than acceptors. Reduced intraclutch variation may enable the host to discriminate a parasitic egg by standardising its own eggs so that it is easier to separate even a partially mimetic parasitic egg from their own clutch. Human assessment of photographs did not reveal any significant effect of contrast of the introduced egg on rejection behaviour. In the present study, test persons assessed the eggs both as a whole and each egg part separately. However, many introduced eggs scored as nonmimetic by the human observers were accepted by the hosts. This result is contrary to the one obtained in a previous study on blackcaps from the same area (Honza
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Table 2. Models of egg rejection behaviour dependent on characteristics of egg colour in the blackcap using partial regression coefficients and other statistics of multiple logistic regression Variable
Model 1
Model 2
Model 3
Model 4
Model 5
Model 6
Model 7
Intercept b1(Chroma UV intraclutch variation) IMD b2(Chroma blue intraclutch variation) IMD b3(Chroma green intraclutch variation) IMD b4(Chroma red intraclutch variation) IMD b5(Blue whole egg) IMD b6(Green whole egg) IMD b7(UV blunt egg part) IMD b8(Blue blunt egg part) IMD b9(Green blunt egg part) IMD b10(Red blunt egg part) IMD b11(Yellow sharp egg part) IMD b12(Red sharp egg part) IMD b13(Brightness blunt egg part) IMD
1.76**** d d d d d d d d d d d d 4.88**** 12.35**** d d d d d d d d d d d d
3.14**** d d 3.71*** 4.08**** d d d d d d d d 4.83**** 10.84**** d d d d d d d d d d d d
5.46**** d d 4.91**** 5.82**** d d d d d d d d 4.54**** 7.80**** d d d d d d d d d d 5.16*** 4.18****
6.08**** d d 9.21**** 8.47**** d d 4.77* 2.85*** d d d d 4.56**** 6.69**** d d d d d d d d d d 6.20*** 4.68****
3.20**** d d 7.14** 5.62**** 6.47*** 5.58**** 9.20*** 7.33**** d d d d 8.42**** 15.38**** d d d d d d d d 6.74**** 5.54**** d d
8.90**** d d 16.34**** 11.73**** 8.15**** 7.00**** 13.80*** 9.30**** d d d d 8.95**** 12.36**** d d d d d d 6.33*** 4.61**** d d 10.32**** 7.36****
8.09**** 8.00**** 6.89**** d d d d d d 7.87** 2.91*** 10.10*** 7.08**** d d 19.67**** 15.15**** 18.75**** 11.57**** NS 1.69* d d d d 8.20**** 6.32****
Model deviance Increase from model deviance Model R2 % Correctly classified total Area under ROC curve 2
45.33 12.35 0.22 72.73 0.801
41.25 16.43 0.28 86.36 0.863
37.07 20.61 0.36 77.27 0.871
34.22 23.46 0.41 79.54 0.887
30.75 26.93 0.47 84.09 0.926
25.68 32.00 0.56 86.36 0.948
25.39 32.29 0.56 90.91 0.936
For details see Methods. b1eb9, The estimates of partial regression coefficients; IMD, increase from model deviance (the change in deviance due to excluding corresponding independent variable from the model); Wald test: ****P value < 0.05; ***0.05 < P < 0.1; **0.1 < P < 0.15; *0.15 < P < 0.2; NS, P > 0.2; d variable not included in the model by hierarchical forward subset selection.
et al. 2004). This can be caused by lower differences in contrast between acceptors and rejecters in the present study (acceptors 2.18 0.48, rejecters 2.40 0.54) compared to the previous study (acceptors 1.66 0.48, rejecters 2.25 0.59). It is difficult to explain this discrepancy, but we suppose that this might be due to different number of
45 Reflectance (%)
424
40 35 30 25 20 15 10 300
400
500
600
700
Wavelength (nm) Figure 1. Mean reflectance spectra of the blunt egg part in blackcap clutches, where parasitic egg was accepted and rejected. Acceptors N1 ¼ 28 (upper line), rejecters N2 ¼ 16 (lower line).
the persons who assessed the clutches (10 and three, respectively). Similarly, we did not confirm the relationship between rejection and contrast when we tested mean values of reflectance measurements per egg. Nevertheless, models derived from a more detailed spectrophotometric analysis showed that egg rejection depends strongly and consistently only on contrast of the blunt part in UV spectra and brightness. These two variables tended to correlate, suggesting that decrease of contrast in UV spectra might be associated with decreasing brightness of the blunt part. Cherry & Bennett (2001) emphasized the importance of UV part of spectrum for the discrimination of cuckoo egg by the Cape robin-chat, Cossypha caffra. Similarly, Avile´s & Møller (2003) confirmed the importance of the part of the spectrum invisible to humans. They found that allopatric population of meadow pipits, Anthus pratensis, showed greater variation within clutches in egg appearance at the ultraviolet wavelengths than a population sympatric with common cuckoo. Similarly, Soler et al. (2003) found that great spotted cuckoo eggs differed from host eggs not only in other parts of the light spectrum but also in UV. In our study we also confirmed that not only the part of the
ET AL.: EGG RECOGNITION IN BLACKCAPS POLA CIKOV A
spectrum visible to humans is important in rejection of foreign eggs, but their combination with UV wavelengths plays a significant role in egg discrimination of blackcap. As stated above, the regression models also showed that the probability of rejection is significantly influenced by the brightness of the blunt egg part. The darker the blunt egg part of the host clutch was, the more probable the rejection. As with UV reflectance, this finding highlights the importance of coloration of the blunt egg pole. Bischoff & Murphy (1993) compared within-clutch variation in the eastern kingbird, Tyrannus tyrannus, and found that egg coloration of different females varied considerably in eggshell markings only at the upper third of the egg. A similar result was reported by Brown & Sherman (1989) who measured variation in egg appearance within and between clutches of barn swallows, Hirundo rustica, and cliff swallows, Hirundo pyrrhonota. For both species they found that within-clutch variance was significantly lower than between-clutch variance for some variables including spotting at the blunt end of the egg. It is generally expected that rejection rate of parasitic eggs should increase with increasing values of contrast between host and parasitic eggs (Davies & de Brooke 1989), but as stated above, we found the opposite. The likelihood of egg rejection increased with decreasing values of absolute differences of mean reflectance in the blunt part at UV wavelengths between host and parasitic eggs. Since we showed that brightness of the blunt egg part was lower in rejecters than acceptors, we compared the brightness of acceptors and rejecters clutches versus accepted and rejected foreign eggs in their blunt part of eggs. Accepted eggs were significantly darker, whereas rejected eggs were somewhat lighter in their blunt parts compared to the host clutches. Our counterintuitive result that the probability of foreign egg acceptance increases with increasing contrast between host and parasitic eggs might be linked with an underlying direct relationship (the darker the parasitic egg, the higher its acceptance rate). Multivariate perspective of our models enables us to propose that rejection behaviour towards alien eggs does not depend on contrast values alone, but with equal strength also on differences in brightness between host and parasitic egg. If birds reject every egg even a little bit lighter than host eggs and accept all eggs which are as dark as or much darker than the host clutch, then contrast between host and foreign eggs will decrease in rejecters to the degree in which rejecters’ clutches are darker than acceptors’. Moreover, in rejecters with darker and more uniform clutches and apparently better discriminating abilities even little contrast in alien egg appearance might be sufficient for egg recognition, whereas in acceptors much greater contrasts are likely to go unnoticed. Although it is difficult to explain why eggs darker than the host eggs were tolerated in the nest and vice versa, this pattern may be connected with the fact that darker eggs of the rejecters may have something to do with the concentration of the spots at the blunt egg part in the blackcap (Makatsch 1976), which might theoretically serve as a fingerprint of individual females (Kilner 2006). Alternatively, if synthesis and allocation of pigments into the eggshell is costly (Moreno & Osorno 2003), the darkness of the blunt egg
part may be linked to the quality and age of the bird (Moreno et al. 2006; Siefferman et al. 2006), which can be further reflected in the host rejection response (Lotem et al. 1995). Unfortunately, we have no data on the age and quality of our blackcaps and thus this is rather a speculation and a challenge to future studies. Methods based on human vision and on mean values of spectrophotometric measurements per individual egg did not support our prediction that contrast between blackcap and parasitic eggs influences egg rejection. Previous spectrophotometric studies on egg coloration (Cherry & Bennett 2001; Soler et al. 2003, 2005; Avile´s & Møller 2004) found a high repeatability among spectrometric measurements on host and parasitic eggs and they subsequently used only mean values for each egg. Because of the complexity of the colour pattern on the eggshell, we analysed three parts of the eggshell separately and showed that some differences within the eggshell may remain undetected when calculating averages for egg as a whole. Our data provide some evidence that the blunt pole of passerine eggs is the most variable part of their eggs and may thus serve as an important cue for egg recognition. This result has methodological implications for the assessment of egg appearance in brood parasitic studies. To conclude, human assessment of individual egg parts and mean values of spectrophotometric measurements per individual egg did not reveal the factors responsible for egg discrimination. However, by using spectrophotometric analyses of separate parts of the egg we showed that egg recognition in blackcaps is significantly influenced by intraclutch variation, the degree of mimicry of the parasitic egg and the brightness of the blunt egg parts. Therefore, this study revealed that particular egg parts may play a role in egg rejection and more detailed spectrophotometric studies are needed to explore a possible role of the blunt part of the eggs as a cue for recognition. Acknowledgments L.P. supplemented design of the experiment, which had been suggested by M.H. L.P. ran experiments, processed and partly analysed data and wrote the manuscript. L.P.’s fieldwork was covered by the project no. 524/05/H536. M.H. financially supported the experiment by the projects (grant no. A600930605, LC0603). P.P. ran the experiment, discussed and commented on the data analysis. J.T. analysed most of the data. B.G.S. helped with the method of measurements and structuring of the manuscript. B.G.S. was funded by Torstein Erbos Gavefond. All coauthors commented on the manuscript. Thanks are due to M. Polacik, M. Pozgayova´, and V. Sˇicha for various contributions to this work, and M. Dusˇkova´, P. Ha´jkova´, M. Jana´c, M. Ondrackova´, Z. Valova´, P. Vallo, V. Vranova´ and J. Sˇvanyga for assessing the photographs of the clutches. References Antonov, A., Stokke, B. G., Moksnes, A. & Røskaft, E. 2006. Egg rejection in a marsh warbler (Acrocephalus palustris) population
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heavily parasitized by common cuckoos (Cuculus canorus). Auk, 123, 419e430. Avile´s, J. M. & Møller, A. P. 2003. Meadow pipit (Anthus pratensis) egg appearance in cuckoo (Cuculus canorus) sympatric and allopatric populations. Biological Journal of the Linnean Society, 79, 543e549. Avile´s, J. M. & Møller, A. P. 2004. How is host egg mimicry maintained in the cuckoo (Cuculus canorus). Biological Journal of the Linnean Society, 82, 57e68. Avile´s, J. M., Soler, J. J., Pe´rez-Contreras, T., Soler, M. & Møller, A. P. 2006. Ultraviolet reflectance of great spotted cuckoo eggs and egg discrimination by magpies. Behavioral Ecology, 17, 310e314. Bennett, A. T. D. & Cuthill, I. C. 1994. Ultraviolet vision in birds: what is the function? Vision Research, 34, 1471e1478. Bennett, A. T. D., Cuthill, I. C., Partridge, J. C. & Lunau, K. 1997. Ultraviolet plumage colors predict mate preferences in starlings. Proceedings of the National Academy of Sciences, U.S.A., 94, 8618e8621. Berthold, P., Fiedler, W. & Querner, U. 1995. Die Mo ¨ nchsgrasmu¨cke (Sylvia atricapilla) als Kuckucks (Cuculus canorus) Wirt. Charadrius, 31, 11e18. Birkhead, T. R. 1978. Behavioural adaptations to high density nesting in the common guillemot (Uria aalge). Animal Behaviour, 26, 321e331. Bischoff, C. M. & Murphy, M. T. 1993. The detection of and responses to experimental intraspecific brood parasitism in eastern kingbirds. Animal Behaviour, 45, 631e638. Bowmaker, J. K., Heath, L. A., Wilkie, S. E. & Hunt, D. M. 1997. Visual pigments and oil droplets from six classes of photoreceptor in the retinas of birds. Vision Research, 37, 2183e2194. Braa, A. T., Moksnes, A. & Røskaft, E. 1992. Adaptations of bramblings and chaffinches towards parasitism by the common cuckoo. Animal Behaviour, 43, 67e78. Brown, C. R. & Sherman, L. C. 1989. Variation in the appearance of swallow eggs and the detection of intraspecific brood parasitism. Condor, 91, 620e627. Cherry, M. I. & Bennett, A. T. D. 2001. Egg colour matching in an African cuckoo, as revealed by ultravioletevisible reflectance spectrophotometry. Proceedings of the Royal Society of London, Series B, 268, 565e571. Cuthill, I. C., Bennett, A. T. D., Partridge, J. C. & Maier, E. J. 1999. Plumage reflectance and the objective assessment of avian sexual dichromatism. American Naturalist, 153, 183e200. Davies, N. B. 2000. Cuckoos, Cowbirds and Other Cheats. London: T. & A. D. Poyser. Davies, N. B. & de Brooke, M. L. 1989. An experimental study of co-evolution between the cuckoo Cuculus canorus, and its hosts. I. Host egg discrimination. Journal of Animal Ecology, 58, 207e224. Dawkins, R. & Krebs, J. R. 1979. Arms races between and within species. Proceedings of the Royal Society of London, Series B, 205, 489e511. Endler, J. A. 1990. On the measurement and classification of colour in studies of animal colour patterns. Biological Journal of the Linnean Society, 41, 315e352. Goldsmith, T. H. 1991. The evolution of visual pigments and colour vision. In: The Perception of Colour (Ed. by P. Gouras), pp. 62e89. London: Macmillan. Gosler, A. G., Barnett, P. R. & Reynolds, J. 2000. Inheritance and variation in eggshell patterning in the great tit Parus major. Proceedings of the Royal Society of London, Series B, 267, 2469e 2473. Hays, H. & LeCroy, M. 1971. Field criteria for determining incubation stages in eggs of the common tern. Wilson Bulletin, 83, 425e429. Hintze, J. L. 1997e2004. NCSS Statistical System. User’s Guide IeV. Kaysville: Number Cruncher Statistical Systems.
Honkavaara, J., Koivula, M., Korpima¨ki, E., Siitari, H. & Viitala, J. 2002. Ultraviolet vision and foraging in terrestrial vertebrates. Oikos, 98, 505e511. Honza, M., Moksnes, A., Røskaft, E. & Stokke, B. G. 2001. How are different common cuckoo Cuculus canorus egg morphs maintained? An evaluation of different hypotheses. Ardea, 89, 341e352. Honza, M., Procha´zka, P., Stokke, B. G., Moksnes, A., Røskaft, E., Capek, M., Jr & Mrlı´k, V. 2004. Are blackcaps current winners in the evolutionary struggle against the common cuckoo? Journal of Ethology, 22, 175e180. Hoyt, D. F. 1979. Practical methods of estimating volume and fresh weight of bird eggs. Auk, 96, 73e77. Kilner, R. M. 2006. The evolution of egg colour and patterning in birds. Biological Reviews, 81, 383e406. Lack, D. 1968. Ecological Adaptations for Breeding in Birds. London: Methuen. Legendre, L. & Legendre, P. 1983. Numerical Ecology. Developments in Environmental Modelling Vol. 3. Amsterdam: Elsevier Science. Lotem, A., Nakamura, H. & Zahavi, A. 1995. Constraints on egg discrimination and cuckoo-host co-evolution. Animal Behaviour, 49, 1185e1209. Makatsch, W. 1976. Die Eier der Vo¨gel Europas. Radebeul: Neumann. Moksnes, A. & Røskaft, E. 1992. Responses of some rare cuckoo hosts to mimetic model cuckoo eggs and to foreign conspecific eggs. Ornis Scandinavica, 23, 17e23. Moksnes, A. & Røskaft, E. 1995. Egg-morphs and host preference in the common cuckoo (Cuculus canorus): an analysis of cuckoo and host eggs from European museum collections. Journal of Zoology (London), 236, 625e648. Moksnes, A., Røskaft, E., Braa, A. T., Korsnes, L., Lampe, H. M. & Pedersen, H. C. 1990. Behavioural responses of potential hosts towards artificial cuckoo eggs and dummies. Behaviour, 116, 64e89. Moreno, J. & Osorno, J. L. 2003. Avian egg colour and sexual selection: does eggshell pigmentation reflect female condition and genetic quality? Ecology Letters, 6, 803e806. Moreno, J., Lobato, E., Morales, J., Merino, S., Tomas, G., Martinez-de la Puente, J., Sanz, J. J., Mateo, R. & Soler, J. J. 2006. Experimental evidence that egg color indicates female condition at laying in a songbird. Behavioral Ecology, 17, 651e655. Øien, I. J., Moksnes, A. & Røskaft, E. 1995. Evolution of variation in egg color and marking pattern in European passerines: adaptations in a coevolutionary arms race with the cuckoo, Cuculus canorus. Behavioral Ecology, 6, 166e174. Payne, R. B. 1997. Avian brood parasitism. In: HosteParasite Evolution: General Principles and Avian Models (Ed. by D. H. Clayton & J. Moore), pp. 338e369. Oxford: Oxford University Press. Procha´zka, P. & Honza, M. 2003. Do common whitethroats (Sylvia communis) discriminate against alien eggs? Journal fu¨r Ornithologie, 144, 354e363. Procha´zka, P. & Honza, M. 2004. Egg discrimination in the yellowhammer. Condor, 106, 405e410. Siefferman, L., Navara, K. J. & Hill, G. E. 2006. Egg coloration is correlated with female condition in eastern bluebirds (Sialia sialis). Behavioral Ecology and Sociobiology, 59, 651e656. Sokal, R. R. & Rohlf, F. J. 1997. Biometry. The Principles and Practice of Statistics in Biological Research. 3rd edn. New York: W.H. Freeman. Soler, J. J., Soler, M. & Møller, A. P. 2000. Host recognition of parasite eggs and the physical appearance of host eggs: the magpie and its brood parasite the great spotted cuckoo. Etologı´a, 8, 9e16.
ET AL.: EGG RECOGNITION IN BLACKCAPS POLA CIKOV A
Soler, J. J., Aviles, J. M., Soler, M. & Møller, A. P. 2003. Evolution of host egg mimicry in a brood parasite, the great spotted cuckoo. Biological Journal of the Linnean Society, 79, 551e563. Soler, J. J., Moreno, J., Avile´s, J. M. & Møller, A. P. 2005. Blue and green egg-color intensity is associated with parental effort and mating system in passerines: support for the sexual selection hypothesis. Evolution, 59, 636e644. Solı´s, J. C. & de Lope, F. 1995. Nest and egg crypsis in the groundnesting stone curlew Burhinus oedicnemus. Journal of Avian Biology, 26, 135e138. Stokke, B. G., Moksnes, A., Røskaft, E., Rudolfsen, G. & Honza, M. 1999. Rejection of artificial cuckoo (Cuculus canorus) eggs in relation to variation in egg appearance among reed warblers (Acrocephalus scirpaceus). Proceedings of the Royal Society of London, Series B, 266, 1483e1488. Stokke, B. G., Moksnes, A. & Røskaft, E. 2002. Obligate brood parasites as selective agents for evolution of egg appearance in passerine birds. Evolution, 56, 199e205. Stokke, B. G., Rudolfsen, G., Moksnes, A. & Røskaft, E. 2004. Rejection of conspecific eggs in chaffinches: the effect of age and clutch characteristics. Ethology, 110, 459e470.
Stokke, B. G., Moksnes, A. & Røskaft, E. 2005. The enigma of imperfect adaptations in hosts of avian brood parasites. Ornithological Science, 4, 17e29. Underwood, T. J. 2003. Proximate and ultimate influences on egg recognition and rejection behavior in response to avian brood parasitism. Ph.D. thesis, University of Manitoba, Winnipeg, Canada. Underwood, T. J. & Sealy, S. G. 2002. Adaptive significance of egg coloration. In: Avian Incubation, Behaviour, Environment and Evolution (Ed. by D. C. Deeming), pp. 280e298. Oxford: Oxford University Press. Vorobyev, M., Osorio, D., Bennett, A. T. D., Marshall, N. J. & Cuthill, I. C. 1998. Tetrachromacy, oil droplets and bird plumage colours. Journal of Comparative Physiology A, 183, 621e633. Welbergen, J., Komdeur, J., Kats, R. & Berg, M. 2001. Egg discrimination in the Australian reed warbler (Acrocephalus australis): rejection response toward model and conspecific eggs depending on timing and mode of artificial parasitism. Behavioral Ecology, 12, 8e15. Zar, J. H. 1996. Biostatistical Analysis. 3rd edn. Upper Saddle River, New Jersey: Prentice Hall.
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Colour characteristics of the blunt egg pole: cues for recognition of parasitic eggs as revealed by reflectance spectrophotometry OVA ´ *†, M ARC EL H ONZ A* , PETR P ROCH A ´ ZK A* , JA ´ N TOP ERCER‡ & B A˚ R D G. ST OKK E§ LENKA POLA CIK
*Institute of Vertebrate Biology, v.v.i, Academy of Sciences of the Czech Republic yInstitute of Botany and Zoology, Masaryk University in Brno zBotanical Garden, Comenius University xDepartment of Biology, Norwegian University of Science and Technology (Received 12 May 2006; initial acceptance 28 June 2006; final acceptance 16 October 2006; published online 22 August 2007; MS. number: 8958R)
Variation in appearance of parasitic and host eggs is an essential trait in the coevolutionary struggle between a brood parasite and its host. Here we examined the role of colour characteristics in the egg recognition in blackcaps, Sylvia atricapilla, assessed by (1) human vision and (2) spectrophotometry, including UV wavelengths. Since colour and spotting pattern within eggs are often highly variable, we also assessed three different parts of blackcap eggshell separately. We investigated the possibility that a lower intraclutch variation in the parasitized clutch would positively affect egg recognition. In addition, we predicted that egg rejection would depend on contrast between host and parasitic eggs. However, we found that neither intraclutch variation nor mimicry if assessed by humans had significant effect on host rejection. Multivariate analyses of spectrophotometric data revealed, however, that the likelihood of egg rejection significantly increased with decreasing intraclutch variation in blue chroma. Furthermore, probability of egg rejection significantly increased with decreasing contrast between parasitic and host eggs in UV wavelengths and with decreasing brightness of host eggs at the blunt egg pole. Accepted foreign eggs were significantly darker at the blunt pole than acceptors’ own eggs. The paradox that rejected eggs had lower contrast could be explained by the fact that the contrast in UV spectra is associated with brightness of the blunt egg part. Our data suggest that colour characteristics of the blunt egg part may play a major role in the recognition of parasitic eggs in the blackcaps. Ó 2007 The Association for the Study of Animal Behaviour. Published by Elsevier Ltd. All rights reserved.
Keywords: blackcap; brood parasitism; recognition cues; reflectance spectrophotometry; rejection behaviour; Sylvia atricapilla
Various aspects of bird life histories affect the colour and spotting pattern of eggshells (Kilner 2006). Coloration of the bird eggshell varies between and within species, ranging from achromatic pale to diversely spotted eggs (Makatsch 1976; Gosler et al. 2000). Recently, Underwood & Sealy (2002) and Kilner (2006) reviewed several Correspondence: L. Polacikova´, Institute of Vertebrate Biology, v.v.i, Academy of Sciences of the Czech Republic, Kveˇtna´ 8, CZ-603 65 Brno, Czech Republic and the Institute of Botany and Zoology, Masaryk University in Brno, Kotla´ˇrska´ 2, CZ-611 19 Brno, Czech Republic (email: [email protected]). J. Topercer is at the Botanical Garden, Comenius University, SK-038 15 Blatnica, Slovakia. B. G. Stokke is at the Department of Biology, Norwegian University of Science and Technology, NTNU, NO-7491 Trondheim, Norway. 0003e 3472/07/$30.00/0
hypotheses explaining the significance of variability in the colours and patterns on bird eggs. For example, egg coloration may allow clutches to mimic the environment and thus reduce the risk of nest predation (Solı´s & de Lope 1995). The egg colour pattern may also facilitate birds identifying their own clutches in colonially nesting birds (Birkhead 1978). Another notable functional explanation of avian egg coloration is related to the evolution of defences against brood parasitism, allowing hosts to recognise and eject foreign eggs in own clutches. On the other hand, brood parasite may evolve eggs that mimic those of their hosts in both ground colour and spotting pattern (Davies 2000). Brood parasitism in birds provides one of the best models for studying coevolutionary processes consisting
419 Ó 2007 The Association for the Study of Animal Behaviour. Published by Elsevier Ltd. All rights reserved.
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of the reciprocal defences and counterdefences which lead to a coevolutionary ‘arms race’ between hosts and brood parasites (Dawkins & Krebs 1979; Davies & de Brooke 1989). An interspecific brood parasitic reproductive strategy is deployed by about 1% of all bird species (Payne 1997), and comprises laying eggs in nests of another species, the hosts, which incubate the eggs and feed the parasitic young (Lack 1968). If the parasitic act is successful, it generally has detrimental effect on the hosts’ fitness because their reproductive success will be close to zero. This phenomenon represents a strong evolutionary selective force for the hosts to evolve counterdefences enabling them to overcome the negative effects of the parasite (Davies 2000). Several host adaptations related to egg appearance facilitate recognition of the parasitic egg. It has been shown that lower intraclutch and higher interclutch variation in appearance of host eggs (Øien et al. 1995; Stokke et al. 1999, 2002) and especially the contrast between host and parasitic eggs (Davies & de Brooke 1989; Moksnes et al. 1990; Honza et al. 2004; Stokke et al. 2004; Antonov et al. 2006) are the most relevant predictors for acceptance or rejection of foreign eggs. Some previous studies investigated the host ability to assess variation within own clutches based on human perception directly (Stokke et al. 1999; Welbergen et al. 2001; Procha´zka & Honza 2004) or based on image analysis software in the human visible spectrum (Soler et al. 2000). However, several studies assessed egg similarity by reflectance spectrophotometry, because there are anatomical differences between avian and human eyes (Bowmaker et al. 1997; Vorobyev et al. 1998; Cuthill et al. 1999). Birds differ from humans in colour perception (Bennett & Cuthill 1994), their cones being sensitive to a wider spectral range, which enables them perception of wavelengths below 400 nm (Goldsmith 1991). Indeed, recent studies have shown that birds use their ability to perceive ultraviolet wavelengths in mate choice (Bennett et al. 1997; Cuthill et al. 1999) and foraging (Honkavaara et al. 2002). Spectrophotometry techniques have also been applied in studies of avian brood parasitism, and several authors have pointed out the role of UV reflectance in egg recognition (Cherry & Bennett 2001; Soler et al. 2003; Underwood 2003; Avile´s & Møller 2004). However, Avile´s et al. (2006) found that artificial reduction of reflectance in the UV region of great spotted cuckoo, Clamator glandarius, eggs did not affect the ejection of parasitic eggs by their magpie, Pica pica, hosts. In this study, we tested the effect of colour characteristics on parasitic egg rejection in the blackcap, Sylvia atricapilla. This species is a good candidate for such kind of study because its eggs are apparently (based on human vision) highly variable in ground colour and marking pattern between clutches with differently coloured spots, speckles and blotches, which occasionally concentrate on the blunt pole (Makatsch 1976). Furthermore, there is evidence that blackcaps have been used as hosts by the common cuckoo, Cuculus canorus, in the past (Berthold et al. 1995; Moksnes & Røskaft 1995; Honza et al. 2001). As a consequence of this, blackcaps have retained good recognition ability (Moksnes & Røskaft 1992; Honza et al. 2004), and
generally have a low intraclutch and high interclutch variation in egg appearance (Øien et al. 1995). Honza et al. (2004) in a previous study showed that intraclutch variation did not influence rejection responses of the blackcaps. In the light of the above facts, we put forward two predictions. Since this species shows low intraclutch variation, we hypothesized that the intraclutch variation in this species would not significantly affect egg rejection in the hosts as found by Honza et al. (2004). Moreover, we also predicted that egg rejection would depend on contrast between host and parasitic eggs. To test the predictions, we studied the effect of variation in egg appearance within host clutches and degree of mimicry of the introduced egg on rejection behaviour by a traditional method based on human vision and by an objective spectrophotometry analysis. It has already been documented by traditional subjective methods that blackcap rejection behaviour is affected by specific colour characteristics (Honza et al. 2004). However, more objective spectrophotometric techniques can reveal in more detail which part of the light spectrum is important for rejection of foreign eggs. Furthermore, even though the avian egg has a complex colour pattern, previous spectrophotometric studies used mean values of reflectance per egg as a whole. To overcome this, we analysed both mean values per egg and three separate parts of the eggshell (sharp, medium, and blunt part) to test whether certain regions of the egg possess important cues for egg recognition that could be hidden if reflectance from all parts of the egg was averaged. METHODS
Experimental Procedure The study was carried out in a deciduous forest near Dolnı´ Bojanovice (48 510 N, 17 020 E), Czech Republic from 30 April to 22 May 2005. Blackcaps and cuckoos live in sympatry in this area, but we did not record any case of interspecific parasitism during our study. We searched systematically for nests during laying period or the first days of incubation. Eggs found during incubation were floated to estimate their approximate laying date according to methods described by Hays & LeCroy (1971). The eggs were measured (length, L; breadth, B) to the nearest 0.1 mm using callipers. Egg volume (V) was calculated using the formula V ¼ 0.51 L B2 (Hoyt 1979). All eggs were marked by small numbers on the medium part of the eggshell using waterproof ink to allow individual identification. No host egg was rejected, therefore is unlikely that the numbers had any significant effect on egg recognition by the hosts. Blackcap clutches were artificially parasitized either by adding a conspecific egg to the clutch without manipulating the original number of host eggs or exchanging one host egg with a conspecific egg. Altogether 44 experiments were carried out. Conspecific eggs are ideal for their use in this type of research because they are highly variable, ranging from good to poor mimicry. In addition, conspecific eggs are more suitable than model eggs because they are real (models are often deserted), variable in coloration, have natural range of coloration including
ET AL.: EGG RECOGNITION IN BLACKCAPS POLA CIKOV A
in UV, which would be extremely difficult if impossible to mimic using artificial colours. In total, 22 eggs were added and 22 eggs were exchanged during laying period only when at least the fourth host egg was laid. The nests were monitored daily for 5 days after manipulation. If the parasitic egg disappeared within this period but the host eggs were left unharmed, it was considered to have been rejected. If the parasitic egg remained unharmed in the nest, it was considered to have been accepted. If the clutch was abandoned within the 5-day observation period, it was considered to have been deserted. Three depredated clutches were excluded from the analyses.
Estimation of Intraclutch Variation and Contrast from Photographs The whole clutch including the parasitic egg was photographed on a grey background card with KODAK colour scale using a Nikon F65 camera and Fujicolor 200 ASA film. The intraclutch variation in egg appearance and the contrast between parasitic and host eggs were later assessed from the photos by 10 test persons, who judged both whole eggs and each egg part (blunt, medium and sharp) separately (for estimate of contrast), and were unaware of the fate of the introduced eggs. The intraclutch variation in egg ground colour and spotting pattern was scored on the following scale from one to five (Øien et al. 1995). (1) No variation, all the eggs were similar. (2) At least one egg differed slightly from the others. (3) At least one egg showed marked differences from the other eggs. (4) At least one egg differed dramatically from the others. (5) No two eggs were similar. The contrast between the parasitic and host eggs was scored on the following scale from one to three (Braa et al. 1992). (1) No contrast between host and parasitic eggs. The parasitic egg was indistinguishable from the host eggs. (2) Medium contrast between host and parasitic eggs. The parasitic egg could be distinguished from the host eggs, but the difference was only moderate. (3) High contrast between host and parasitic eggs. The parasitic egg could easily be distinguished from the host eggs. The median of scores by the test persons who assessed the whole eggs and the egg parts was used as estimate of intraclutch variation and contrast for each clutch. There was significant repeatability in scoring of the whole eggs among the 10 observers in intraclutch variability (Kendall coefficient of concordance: W43 ¼ 0.55, P < 0.0001) and in contrast between host and foreign eggs (W43 ¼ 0.56, P < 0.0001). Similarly, there was significant repeatability in scoring the contrast between host and parasitic eggs among the 10 observers for the blunt egg parts (W43 ¼ 0.69, P < 0.0001), the medium egg parts (W43 ¼ 0.60, P < 0.000) and the sharp egg parts (W43 ¼ 0.61, P < 0.0001).
Measurement of the Reflectance Spectra We measured the spectral reflectance of all the eggs from the parasitized clutches in the range 300e700 nm using a USB2000 spectrophotometer (Ocean Optics). We divided
each egg into three regions across the longitudinal axis of the egg. Each region comprised a third of the length of the egg representing the blunt, medium and sharp parts of the egg. From each egg part three randomly located measurements were taken (each covering 1 mm2). The illuminant was a deuterium and halogen light source (DT-Mini-GS, Ocean Optics). The light was transferred to the eggs through a quartz optic fibre (QR400-7-UV/VIS-BX, Ocean Optics), reached the eggs and was reflected from the eggs at an angle of 45 to the surface. Data from the spectrophotometer were loaded into OOIBase 32 software (Ocean Optics). The measurements were relative and referred to a standard white reference (WS-2) and to darkness. Reference and dark calibration were made before measurement of each clutch. Total reflectance was obtained for the UV (325e400 nm) interval, which comprises the human invisible part of colour spectrum, and for the blue (400e475 nm), green (475e550 nm), yellow (550e625 nm) and red (625e700 nm) wavelengths as the human visible regions. We calculated objective variables of the colour, namely brightness (reflectance intensity), chroma (purity of a colour) and hue (peak wavelengths) (Endler 1990). Brightness was calculated as the sum of the total reflectance values within the 325e 700 nm range of wavelengths (R325e700). Chroma was calculated as reflectance ratios: R325e400/325e700 (UV chroma), R400e475/325e700 (blue chroma), R475e550/325e700 (green chroma), R550e625/325e700 (yellow chroma) and R625e700/325e700 (red chroma). Hue was estimated by the wavelength of the maximum reflectance (Rmax). We performed the measurements on 193 eggs from 44 clutches. Despite large variation in spottiness of eggs we found significant repeatability for all nine reflectance measurements of each egg (Pearson correlation coefficients: for all 193 eggs: r > 0.80, P < 0.0001). Similarly, we found significant repeatability for all colour variables calculated for each egg (Sokal & Rohlf 1997; for all colour variables P < 0.001, df ¼ 192,1736; Table 1). Therefore we used mean values for each part of the colour spectrum per egg.
Estimation of Intraclutch Variation and Contrast from Spectrophotometric Measurements To estimate the degree of intraclutch variation we calculated coefficient of variation (SD/mean 100) from Table 1. Estimated repeatability (Pearson correlation coefficients) for colour characteristics of nine spectrophotometric measurements per each blackcap egg Colour characteristic Brightness Chroma UV Chroma blue Chroma green Chroma yellow Chroma red Hue
Repeatability
F192,1736
0.24*** 0.32*** 0.37*** 0.28*** 0.58*** 0.37*** 0.53***
3.77 5.25 6.00 4.45 13.5 6.2 11.21
193 Eggs from 44 clutches, ***P < 0.001.
421
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ANIMAL BEHAVIOUR, 74, 3
mean values for the whole egg of all colour variables in each clutch (Avile´s & Møller 2003). To estimate the contrast in coloration between host and parasitic eggs we calculated absolute differences of mean reflectance value in the ultraviolet, blue, green, yellow and red wavelengths among all hosts and parasitic eggs in the same clutch. Similarly, we calculated the absolute differences of mean reflectance values for blunt, medium and sharp part of all hosts and parasitic eggs in the same clutch separately. Mean values of these absolute differences were considered to represent the contrast between host and parasitic eggs. Finally, we compared mean total intensity of light (brightness) reflected from all eggs in each clutch as well as mean reflected light intensity for each egg part of all eggs in each clutch including brightness of accepted and rejected parasitic eggs.
Statistical Analyses To assess differences between the two experimental groups in response to parasitic eggs we used Fisher’s exact test, and for detecting differences in the laying day between acceptors and rejecters we used ManneWhitney U test. Most of the values acquired by scoring the photos (60%) significantly departed from normal distribution (D’AgostinoePearson omnibus test: P < 0.05). Because of this fact and the multivariate nature of the problem, we took advantage of multiple logistic regression with logit link (Hintze 1997e2004; Sokal & Rohlf 1997) to explore simultaneously the effects of intraclutch variation and contrast between host and parasitic eggs and their three parts on rejection behaviour. As all scores were recorded on the ordinal scale of measurement and the arithmetic mean is applicable only to ratio or interval scale data (Zar 1996), we submitted medians instead of means to the analysis. The comparison of reflected light intensity of all eggs in each clutch fully met the assumption of normality (D’AgostinoePearson omnibus test: P > 0.05), whereas the mean values of colour variables of intraclutch variation obtained by spectrophotometry and variables related to differences between parasitic and host eggs in the same nest violated the normality assumption in ca. 71% and 90% of cases, respectively (P < 0.05). Again we used multiple logistic regression to analyse the effects of intraclutch variation and differences in the contrast between host and foreign eggs on rejection behaviour and to assess the relative importance of these effects. To make the effect magnitudes comparable, we converted the values of all colour variables to the same scale of size and variability (from 0 to 1) using transformation proposed by Gower (Legendre & Legendre 1983). We used nonparametric tests for comparing brightness of eggs among egg parts separately (ManneWhitney U test, KruskaleWallis ANOVA test and Bonferroni post hoc test). All analyses proceeded using the statistical systems NCSS 2004 (Hintze 1997e2004) and Statistica 6.0 (StatSoft Inc. 1984e2001, Tulsa, OK, U.S.A.).
Ethical Note Because the eggs were measured under standard light conditions in the laboratory, we exchanged temporarily the host clutch for four to five foreign blackcap eggs to prevent nest desertion. In all nests the birds resumed incubation and none of these eggs was ejected. After the arrival to the lab, eggs were placed in an incubator (Octagon 20, Brinsea, U.K.), with temperature ranging from 37.7 to 37.8 C and relative humidity from 55% to 65%. The eggs were returned to the original nest within 1 h. Twenty-six (59.1%) nests were controlled until hatching. Of these, 14 were predated and 12 survived until hatching and in these nests all eggs hatched. Thus, we are sure that our temporal egg removal had no detrimental effect on hatching success. Moreover, survival to hatching of experimental nests was even slightly higher (Mayfield method, daily survival rate 0.96) compared to a control group of 41 nests (daily survival rate 0.91). All eggs used for experiments were freshly laid and rejection occurred soon after egg addition, that is, in the first days of incubation (incubation period in the blackcap is 12 days). Experiments were conducted in accordance with current laws of the Czech Republic and the Academy of Sciences Animal Care Protocol, licence number: 0008/98-M103.
RESULTS
Experimental Groups In the total of 44 nests, 28 (63.6%) introduced eggs were accepted and 16 (36.4%) eggs were rejected. There were no significant differences in response to alien eggs between the experimental groups in which the parasitic egg was added or exchanged with one of the host eggs (Fisher’s exact test: N1 ¼ 22, N2 ¼ 22, P ¼ 0.75). Also, there was no significant difference in the time to rejection of the parasitic egg between the two experimental groups (Manne Whitney U test: U ¼ 23, N1 ¼ 7, N2 ¼ 9, P ¼ 0.24, median 1 day (interquartile range: IQR 0e2 days), median 1 day (IQR 0e3 days)). Finally, there was no significant difference between the average size of host and the parasitic eggs added to the clutch (mean SD ¼ 2163.51 235.97 mm3, N1 ¼ 101 versus 2454.98 1665.68 mm3, N2 ¼ 22; U ¼ 1017, P ¼ 0.54) or exchanged for one host egg (2106.45 227.61 mm3, N1 ¼ 71 versus 2072.24 244.25 mm3, N2 ¼ 22; U ¼ 672, P ¼ 0.32). Therefore, we pooled the two experimental groups in further analyses.
Intraclutch Variation and Contrast between Host and Parasitic Eggs Assessed by Humans Of the five variables entered into the initial logistic regression model (medians of intraclutch variation and contrast in whole egg and medians of contrasts in three egg parts), none indicated significant effect on reaction (reaction ¼ 2, i.e. rejection) (c21 ¼ 0.37, P ¼ 0.544). Even the best model, containing medians of contrast in whole egg and in the middle part as predictor variables, was not significant (deviance ¼ 56.49, c22 ¼ 2.22, P ¼ 0.33).
ET AL.: EGG RECOGNITION IN BLACKCAPS POLA CIKOV A
Intraclutch Variation and Contrast between Host and Parasitic Eggs Based on Spectrophotometry Of the 31 candidate colour variables, we omitted 14 due to multicollinearity. The rest entered the hierarchical forward subset selection of variables up to one-way interactions with switching (Hintze 1997e2004). We used it in a stepwise manner, that is, we began with the best-fit model with one variable (best single predictor) then built the best-fit model with two variables and so on. Seven logistic regression models were fitted with 13 variables in total (see Table 2), while remaining four variables did not contribute significantly to any model (contrast in coloration of medium egg part at UV and yellow wavelengths, contrast in sharp egg part at blue wavelengths and average brightness; deviance tests: P > 0.2). Each model proved to be highly significant and valid except for that with six variables, in which maximum likelihood algorithm failed to converge and which was thus replaced by the second model. Except for contrast in blunt part at red wavelengths and average contrast at blue wavelengths, each variable showed significant effect on egg rejection behaviour in at least one model (deviance tests: P < 0.05). Contrast in coloration of blunt part at UV wavelengths (also the best single predictor: c21 ¼ 12.35, P ¼ 0.0004), intraclutch variation in blue chroma and brightness of blunt part were the most useful variables in prediction, gaining significance in six, five and four valid models, respectively. Their regression coefficients had consistent negative values similar in size ratios across the models, indicating (1) inverse relationships to rejection behaviour (the likelihood of alien egg rejection increases with decreasing values of both contrast in blunt part at UV wavelengths, intraclutch variation in blue chroma and brightness of blunt part) and (2) roughly equal effect magnitudes of the three variables. A trend towards a positive correlation between two of them (contrast in blunt part at UV wavelengths versus brightness of blunt part: Spearman rank correlation: rS ¼ 0.26, N ¼ 44, P ¼ 0.084) arose from additional correlation analysis. The simplest model for estimating the probability of egg rejection in blackcap using the three most productive predictor variables is: p2 ¼
e½5:464:91ðCh Blue ivÞ4:54ðUV epc1Þ5:16ðBrightness epb1Þ ; 1 þ e½5:464:91ðCh Blue ivÞ4:54ðUV epc1Þ5:16ðBrightness epb1Þ
where p2 is the probability of an egg being rejected, e is the base of natural logarithms, Ch_Blue_iv is the intraclutch variation in blue chroma, UV_epc1 is the contrast in coloration of blunt part at UV wavelengths and Brightness_ epb1 is the brightness of blunt part. Model accuracy was 77.27%, suggesting quite good ability to predict the egg rejection behaviour. Its explanatory power, however, was not that high (R2 ¼ 0.36). To explain the negative regression coefficients relating the rejection behaviour to the absolute differences between host and parasitic eggs at the blunt egg part, we looked closer at the brightness of the introduced and host eggs. The multiple logistic regression based on
spectrophotometry showed that the brightness reflected from the blunt egg part was lower in rejecters than acceptors (Fig. 1), a result consistent with two-sample testing (t42 ¼ 2.13, N1 ¼ 28, N2 ¼ 16, P ¼ 0.02; mean SD ¼ 24 174.58 4195.60 versus 27 465.11 5290.10). When we compared brightness of the blunt parts of accepted and rejected parasitic eggs, we did not confirm any significant differences although accepted eggs tended to be darker than rejected eggs (ManneWhitney U test: U ¼ 157, N1 ¼ 28, N2 ¼ 16, P > 0.05; mean SD ¼ 24 253.83 7992.30 versus 26 245.49 4358.75). Third, we compared the brightness of rejected eggs with the host eggs and we found that blunt egg parts of host clutches were darker than rejected eggs although the difference was not significant (U ¼ 98, N1 ¼ 16, N2 ¼ 16, P > 0.05; 24 174.58 4195.61 versus 26 245.49 4358.75). Finally, when we compared the brightness of accepted eggs with the host eggs, we found that blunt egg parts of host clutches were significantly lighter than accepted eggs (U ¼ 252, N1 ¼ 28, N2 ¼ 28, P ¼ 0.02; 27 465.64 5289.61 versus 24 253.83 7992.30). To confirm the differences between individual parts of an egg, we additionally compared also the colour variables of blunt, medium and sharp parts of all eggs. When using Bonferroni test, we found significant differences between all three egg parts in all colour characteristics (for all P < 0.016, N ¼ 579), except that blunt and medium egg parts did not differ significantly in yellow chroma (P ¼ 0.99).
DISCUSSION The blackcap shows exquisite egg recognition ability and rejects nearly 100% of nonmimetic eggs from its nests (Honza et al. 2004). The majority of individuals are therefore apparently able to reject foreign eggs as long as the appearance of foreign eggs is above a specific threshold for egg recognition based on individual’s cognitive abilities (Stokke et al. 2005). In line with our prediction, logistic regression model analysing the photo scores did not reveal significant influence of intraclutch variation on the host response. Similarly, many previous studies found the same pattern (Procha´zka & Honza 2003, 2004; Honza et al. 2004; Stokke et al. 2004). However, interestingly, our models based on spectrometric measurements revealed that the probability of egg rejection is negatively associated with intraclutch variation in host egg appearance in one part of the spectrum. Birds that rejected parasitic eggs showed lower variation in blue chroma than acceptors. Reduced intraclutch variation may enable the host to discriminate a parasitic egg by standardising its own eggs so that it is easier to separate even a partially mimetic parasitic egg from their own clutch. Human assessment of photographs did not reveal any significant effect of contrast of the introduced egg on rejection behaviour. In the present study, test persons assessed the eggs both as a whole and each egg part separately. However, many introduced eggs scored as nonmimetic by the human observers were accepted by the hosts. This result is contrary to the one obtained in a previous study on blackcaps from the same area (Honza
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Table 2. Models of egg rejection behaviour dependent on characteristics of egg colour in the blackcap using partial regression coefficients and other statistics of multiple logistic regression Variable
Model 1
Model 2
Model 3
Model 4
Model 5
Model 6
Model 7
Intercept b1(Chroma UV intraclutch variation) IMD b2(Chroma blue intraclutch variation) IMD b3(Chroma green intraclutch variation) IMD b4(Chroma red intraclutch variation) IMD b5(Blue whole egg) IMD b6(Green whole egg) IMD b7(UV blunt egg part) IMD b8(Blue blunt egg part) IMD b9(Green blunt egg part) IMD b10(Red blunt egg part) IMD b11(Yellow sharp egg part) IMD b12(Red sharp egg part) IMD b13(Brightness blunt egg part) IMD
1.76**** d d d d d d d d d d d d 4.88**** 12.35**** d d d d d d d d d d d d
3.14**** d d 3.71*** 4.08**** d d d d d d d d 4.83**** 10.84**** d d d d d d d d d d d d
5.46**** d d 4.91**** 5.82**** d d d d d d d d 4.54**** 7.80**** d d d d d d d d d d 5.16*** 4.18****
6.08**** d d 9.21**** 8.47**** d d 4.77* 2.85*** d d d d 4.56**** 6.69**** d d d d d d d d d d 6.20*** 4.68****
3.20**** d d 7.14** 5.62**** 6.47*** 5.58**** 9.20*** 7.33**** d d d d 8.42**** 15.38**** d d d d d d d d 6.74**** 5.54**** d d
8.90**** d d 16.34**** 11.73**** 8.15**** 7.00**** 13.80*** 9.30**** d d d d 8.95**** 12.36**** d d d d d d 6.33*** 4.61**** d d 10.32**** 7.36****
8.09**** 8.00**** 6.89**** d d d d d d 7.87** 2.91*** 10.10*** 7.08**** d d 19.67**** 15.15**** 18.75**** 11.57**** NS 1.69* d d d d 8.20**** 6.32****
Model deviance Increase from model deviance Model R2 % Correctly classified total Area under ROC curve 2
45.33 12.35 0.22 72.73 0.801
41.25 16.43 0.28 86.36 0.863
37.07 20.61 0.36 77.27 0.871
34.22 23.46 0.41 79.54 0.887
30.75 26.93 0.47 84.09 0.926
25.68 32.00 0.56 86.36 0.948
25.39 32.29 0.56 90.91 0.936
For details see Methods. b1eb9, The estimates of partial regression coefficients; IMD, increase from model deviance (the change in deviance due to excluding corresponding independent variable from the model); Wald test: ****P value < 0.05; ***0.05 < P < 0.1; **0.1 < P < 0.15; *0.15 < P < 0.2; NS, P > 0.2; d variable not included in the model by hierarchical forward subset selection.
et al. 2004). This can be caused by lower differences in contrast between acceptors and rejecters in the present study (acceptors 2.18 0.48, rejecters 2.40 0.54) compared to the previous study (acceptors 1.66 0.48, rejecters 2.25 0.59). It is difficult to explain this discrepancy, but we suppose that this might be due to different number of
45 Reflectance (%)
424
40 35 30 25 20 15 10 300
400
500
600
700
Wavelength (nm) Figure 1. Mean reflectance spectra of the blunt egg part in blackcap clutches, where parasitic egg was accepted and rejected. Acceptors N1 ¼ 28 (upper line), rejecters N2 ¼ 16 (lower line).
the persons who assessed the clutches (10 and three, respectively). Similarly, we did not confirm the relationship between rejection and contrast when we tested mean values of reflectance measurements per egg. Nevertheless, models derived from a more detailed spectrophotometric analysis showed that egg rejection depends strongly and consistently only on contrast of the blunt part in UV spectra and brightness. These two variables tended to correlate, suggesting that decrease of contrast in UV spectra might be associated with decreasing brightness of the blunt part. Cherry & Bennett (2001) emphasized the importance of UV part of spectrum for the discrimination of cuckoo egg by the Cape robin-chat, Cossypha caffra. Similarly, Avile´s & Møller (2003) confirmed the importance of the part of the spectrum invisible to humans. They found that allopatric population of meadow pipits, Anthus pratensis, showed greater variation within clutches in egg appearance at the ultraviolet wavelengths than a population sympatric with common cuckoo. Similarly, Soler et al. (2003) found that great spotted cuckoo eggs differed from host eggs not only in other parts of the light spectrum but also in UV. In our study we also confirmed that not only the part of the
ET AL.: EGG RECOGNITION IN BLACKCAPS POLA CIKOV A
spectrum visible to humans is important in rejection of foreign eggs, but their combination with UV wavelengths plays a significant role in egg discrimination of blackcap. As stated above, the regression models also showed that the probability of rejection is significantly influenced by the brightness of the blunt egg part. The darker the blunt egg part of the host clutch was, the more probable the rejection. As with UV reflectance, this finding highlights the importance of coloration of the blunt egg pole. Bischoff & Murphy (1993) compared within-clutch variation in the eastern kingbird, Tyrannus tyrannus, and found that egg coloration of different females varied considerably in eggshell markings only at the upper third of the egg. A similar result was reported by Brown & Sherman (1989) who measured variation in egg appearance within and between clutches of barn swallows, Hirundo rustica, and cliff swallows, Hirundo pyrrhonota. For both species they found that within-clutch variance was significantly lower than between-clutch variance for some variables including spotting at the blunt end of the egg. It is generally expected that rejection rate of parasitic eggs should increase with increasing values of contrast between host and parasitic eggs (Davies & de Brooke 1989), but as stated above, we found the opposite. The likelihood of egg rejection increased with decreasing values of absolute differences of mean reflectance in the blunt part at UV wavelengths between host and parasitic eggs. Since we showed that brightness of the blunt egg part was lower in rejecters than acceptors, we compared the brightness of acceptors and rejecters clutches versus accepted and rejected foreign eggs in their blunt part of eggs. Accepted eggs were significantly darker, whereas rejected eggs were somewhat lighter in their blunt parts compared to the host clutches. Our counterintuitive result that the probability of foreign egg acceptance increases with increasing contrast between host and parasitic eggs might be linked with an underlying direct relationship (the darker the parasitic egg, the higher its acceptance rate). Multivariate perspective of our models enables us to propose that rejection behaviour towards alien eggs does not depend on contrast values alone, but with equal strength also on differences in brightness between host and parasitic egg. If birds reject every egg even a little bit lighter than host eggs and accept all eggs which are as dark as or much darker than the host clutch, then contrast between host and foreign eggs will decrease in rejecters to the degree in which rejecters’ clutches are darker than acceptors’. Moreover, in rejecters with darker and more uniform clutches and apparently better discriminating abilities even little contrast in alien egg appearance might be sufficient for egg recognition, whereas in acceptors much greater contrasts are likely to go unnoticed. Although it is difficult to explain why eggs darker than the host eggs were tolerated in the nest and vice versa, this pattern may be connected with the fact that darker eggs of the rejecters may have something to do with the concentration of the spots at the blunt egg part in the blackcap (Makatsch 1976), which might theoretically serve as a fingerprint of individual females (Kilner 2006). Alternatively, if synthesis and allocation of pigments into the eggshell is costly (Moreno & Osorno 2003), the darkness of the blunt egg
part may be linked to the quality and age of the bird (Moreno et al. 2006; Siefferman et al. 2006), which can be further reflected in the host rejection response (Lotem et al. 1995). Unfortunately, we have no data on the age and quality of our blackcaps and thus this is rather a speculation and a challenge to future studies. Methods based on human vision and on mean values of spectrophotometric measurements per individual egg did not support our prediction that contrast between blackcap and parasitic eggs influences egg rejection. Previous spectrophotometric studies on egg coloration (Cherry & Bennett 2001; Soler et al. 2003, 2005; Avile´s & Møller 2004) found a high repeatability among spectrometric measurements on host and parasitic eggs and they subsequently used only mean values for each egg. Because of the complexity of the colour pattern on the eggshell, we analysed three parts of the eggshell separately and showed that some differences within the eggshell may remain undetected when calculating averages for egg as a whole. Our data provide some evidence that the blunt pole of passerine eggs is the most variable part of their eggs and may thus serve as an important cue for egg recognition. This result has methodological implications for the assessment of egg appearance in brood parasitic studies. To conclude, human assessment of individual egg parts and mean values of spectrophotometric measurements per individual egg did not reveal the factors responsible for egg discrimination. However, by using spectrophotometric analyses of separate parts of the egg we showed that egg recognition in blackcaps is significantly influenced by intraclutch variation, the degree of mimicry of the parasitic egg and the brightness of the blunt egg parts. Therefore, this study revealed that particular egg parts may play a role in egg rejection and more detailed spectrophotometric studies are needed to explore a possible role of the blunt part of the eggs as a cue for recognition. Acknowledgments L.P. supplemented design of the experiment, which had been suggested by M.H. L.P. ran experiments, processed and partly analysed data and wrote the manuscript. L.P.’s fieldwork was covered by the project no. 524/05/H536. M.H. financially supported the experiment by the projects (grant no. A600930605, LC0603). P.P. ran the experiment, discussed and commented on the data analysis. J.T. analysed most of the data. B.G.S. helped with the method of measurements and structuring of the manuscript. B.G.S. was funded by Torstein Erbos Gavefond. All coauthors commented on the manuscript. Thanks are due to M. Polacik, M. Pozgayova´, and V. Sˇicha for various contributions to this work, and M. Dusˇkova´, P. Ha´jkova´, M. Jana´c, M. Ondrackova´, Z. Valova´, P. Vallo, V. Vranova´ and J. Sˇvanyga for assessing the photographs of the clutches. References Antonov, A., Stokke, B. G., Moksnes, A. & Røskaft, E. 2006. Egg rejection in a marsh warbler (Acrocephalus palustris) population
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ANIMAL BEHAVIOUR, 74, 3
heavily parasitized by common cuckoos (Cuculus canorus). Auk, 123, 419e430. Avile´s, J. M. & Møller, A. P. 2003. Meadow pipit (Anthus pratensis) egg appearance in cuckoo (Cuculus canorus) sympatric and allopatric populations. Biological Journal of the Linnean Society, 79, 543e549. Avile´s, J. M. & Møller, A. P. 2004. How is host egg mimicry maintained in the cuckoo (Cuculus canorus). Biological Journal of the Linnean Society, 82, 57e68. Avile´s, J. M., Soler, J. J., Pe´rez-Contreras, T., Soler, M. & Møller, A. P. 2006. Ultraviolet reflectance of great spotted cuckoo eggs and egg discrimination by magpies. Behavioral Ecology, 17, 310e314. Bennett, A. T. D. & Cuthill, I. C. 1994. Ultraviolet vision in birds: what is the function? Vision Research, 34, 1471e1478. Bennett, A. T. D., Cuthill, I. C., Partridge, J. C. & Lunau, K. 1997. Ultraviolet plumage colors predict mate preferences in starlings. Proceedings of the National Academy of Sciences, U.S.A., 94, 8618e8621. Berthold, P., Fiedler, W. & Querner, U. 1995. Die Mo ¨ nchsgrasmu¨cke (Sylvia atricapilla) als Kuckucks (Cuculus canorus) Wirt. Charadrius, 31, 11e18. Birkhead, T. R. 1978. Behavioural adaptations to high density nesting in the common guillemot (Uria aalge). Animal Behaviour, 26, 321e331. Bischoff, C. M. & Murphy, M. T. 1993. The detection of and responses to experimental intraspecific brood parasitism in eastern kingbirds. Animal Behaviour, 45, 631e638. Bowmaker, J. K., Heath, L. A., Wilkie, S. E. & Hunt, D. M. 1997. Visual pigments and oil droplets from six classes of photoreceptor in the retinas of birds. Vision Research, 37, 2183e2194. Braa, A. T., Moksnes, A. & Røskaft, E. 1992. Adaptations of bramblings and chaffinches towards parasitism by the common cuckoo. Animal Behaviour, 43, 67e78. Brown, C. R. & Sherman, L. C. 1989. Variation in the appearance of swallow eggs and the detection of intraspecific brood parasitism. Condor, 91, 620e627. Cherry, M. I. & Bennett, A. T. D. 2001. Egg colour matching in an African cuckoo, as revealed by ultravioletevisible reflectance spectrophotometry. Proceedings of the Royal Society of London, Series B, 268, 565e571. Cuthill, I. C., Bennett, A. T. D., Partridge, J. C. & Maier, E. J. 1999. Plumage reflectance and the objective assessment of avian sexual dichromatism. American Naturalist, 153, 183e200. Davies, N. B. 2000. Cuckoos, Cowbirds and Other Cheats. London: T. & A. D. Poyser. Davies, N. B. & de Brooke, M. L. 1989. An experimental study of co-evolution between the cuckoo Cuculus canorus, and its hosts. I. Host egg discrimination. Journal of Animal Ecology, 58, 207e224. Dawkins, R. & Krebs, J. R. 1979. Arms races between and within species. Proceedings of the Royal Society of London, Series B, 205, 489e511. Endler, J. A. 1990. On the measurement and classification of colour in studies of animal colour patterns. Biological Journal of the Linnean Society, 41, 315e352. Goldsmith, T. H. 1991. The evolution of visual pigments and colour vision. In: The Perception of Colour (Ed. by P. Gouras), pp. 62e89. London: Macmillan. Gosler, A. G., Barnett, P. R. & Reynolds, J. 2000. Inheritance and variation in eggshell patterning in the great tit Parus major. Proceedings of the Royal Society of London, Series B, 267, 2469e 2473. Hays, H. & LeCroy, M. 1971. Field criteria for determining incubation stages in eggs of the common tern. Wilson Bulletin, 83, 425e429. Hintze, J. L. 1997e2004. NCSS Statistical System. User’s Guide IeV. Kaysville: Number Cruncher Statistical Systems.
Honkavaara, J., Koivula, M., Korpima¨ki, E., Siitari, H. & Viitala, J. 2002. Ultraviolet vision and foraging in terrestrial vertebrates. Oikos, 98, 505e511. Honza, M., Moksnes, A., Røskaft, E. & Stokke, B. G. 2001. How are different common cuckoo Cuculus canorus egg morphs maintained? An evaluation of different hypotheses. Ardea, 89, 341e352. Honza, M., Procha´zka, P., Stokke, B. G., Moksnes, A., Røskaft, E., Capek, M., Jr & Mrlı´k, V. 2004. Are blackcaps current winners in the evolutionary struggle against the common cuckoo? Journal of Ethology, 22, 175e180. Hoyt, D. F. 1979. Practical methods of estimating volume and fresh weight of bird eggs. Auk, 96, 73e77. Kilner, R. M. 2006. The evolution of egg colour and patterning in birds. Biological Reviews, 81, 383e406. Lack, D. 1968. Ecological Adaptations for Breeding in Birds. London: Methuen. Legendre, L. & Legendre, P. 1983. Numerical Ecology. Developments in Environmental Modelling Vol. 3. Amsterdam: Elsevier Science. Lotem, A., Nakamura, H. & Zahavi, A. 1995. Constraints on egg discrimination and cuckoo-host co-evolution. Animal Behaviour, 49, 1185e1209. Makatsch, W. 1976. Die Eier der Vo¨gel Europas. Radebeul: Neumann. Moksnes, A. & Røskaft, E. 1992. Responses of some rare cuckoo hosts to mimetic model cuckoo eggs and to foreign conspecific eggs. Ornis Scandinavica, 23, 17e23. Moksnes, A. & Røskaft, E. 1995. Egg-morphs and host preference in the common cuckoo (Cuculus canorus): an analysis of cuckoo and host eggs from European museum collections. Journal of Zoology (London), 236, 625e648. Moksnes, A., Røskaft, E., Braa, A. T., Korsnes, L., Lampe, H. M. & Pedersen, H. C. 1990. Behavioural responses of potential hosts towards artificial cuckoo eggs and dummies. Behaviour, 116, 64e89. Moreno, J. & Osorno, J. L. 2003. Avian egg colour and sexual selection: does eggshell pigmentation reflect female condition and genetic quality? Ecology Letters, 6, 803e806. Moreno, J., Lobato, E., Morales, J., Merino, S., Tomas, G., Martinez-de la Puente, J., Sanz, J. J., Mateo, R. & Soler, J. J. 2006. Experimental evidence that egg color indicates female condition at laying in a songbird. Behavioral Ecology, 17, 651e655. Øien, I. J., Moksnes, A. & Røskaft, E. 1995. Evolution of variation in egg color and marking pattern in European passerines: adaptations in a coevolutionary arms race with the cuckoo, Cuculus canorus. Behavioral Ecology, 6, 166e174. Payne, R. B. 1997. Avian brood parasitism. In: HosteParasite Evolution: General Principles and Avian Models (Ed. by D. H. Clayton & J. Moore), pp. 338e369. Oxford: Oxford University Press. Procha´zka, P. & Honza, M. 2003. Do common whitethroats (Sylvia communis) discriminate against alien eggs? Journal fu¨r Ornithologie, 144, 354e363. Procha´zka, P. & Honza, M. 2004. Egg discrimination in the yellowhammer. Condor, 106, 405e410. Siefferman, L., Navara, K. J. & Hill, G. E. 2006. Egg coloration is correlated with female condition in eastern bluebirds (Sialia sialis). Behavioral Ecology and Sociobiology, 59, 651e656. Sokal, R. R. & Rohlf, F. J. 1997. Biometry. The Principles and Practice of Statistics in Biological Research. 3rd edn. New York: W.H. Freeman. Soler, J. J., Soler, M. & Møller, A. P. 2000. Host recognition of parasite eggs and the physical appearance of host eggs: the magpie and its brood parasite the great spotted cuckoo. Etologı´a, 8, 9e16.
ET AL.: EGG RECOGNITION IN BLACKCAPS POLA CIKOV A
Soler, J. J., Aviles, J. M., Soler, M. & Møller, A. P. 2003. Evolution of host egg mimicry in a brood parasite, the great spotted cuckoo. Biological Journal of the Linnean Society, 79, 551e563. Soler, J. J., Moreno, J., Avile´s, J. M. & Møller, A. P. 2005. Blue and green egg-color intensity is associated with parental effort and mating system in passerines: support for the sexual selection hypothesis. Evolution, 59, 636e644. Solı´s, J. C. & de Lope, F. 1995. Nest and egg crypsis in the groundnesting stone curlew Burhinus oedicnemus. Journal of Avian Biology, 26, 135e138. Stokke, B. G., Moksnes, A., Røskaft, E., Rudolfsen, G. & Honza, M. 1999. Rejection of artificial cuckoo (Cuculus canorus) eggs in relation to variation in egg appearance among reed warblers (Acrocephalus scirpaceus). Proceedings of the Royal Society of London, Series B, 266, 1483e1488. Stokke, B. G., Moksnes, A. & Røskaft, E. 2002. Obligate brood parasites as selective agents for evolution of egg appearance in passerine birds. Evolution, 56, 199e205. Stokke, B. G., Rudolfsen, G., Moksnes, A. & Røskaft, E. 2004. Rejection of conspecific eggs in chaffinches: the effect of age and clutch characteristics. Ethology, 110, 459e470.
Stokke, B. G., Moksnes, A. & Røskaft, E. 2005. The enigma of imperfect adaptations in hosts of avian brood parasites. Ornithological Science, 4, 17e29. Underwood, T. J. 2003. Proximate and ultimate influences on egg recognition and rejection behavior in response to avian brood parasitism. Ph.D. thesis, University of Manitoba, Winnipeg, Canada. Underwood, T. J. & Sealy, S. G. 2002. Adaptive significance of egg coloration. In: Avian Incubation, Behaviour, Environment and Evolution (Ed. by D. C. Deeming), pp. 280e298. Oxford: Oxford University Press. Vorobyev, M., Osorio, D., Bennett, A. T. D., Marshall, N. J. & Cuthill, I. C. 1998. Tetrachromacy, oil droplets and bird plumage colours. Journal of Comparative Physiology A, 183, 621e633. Welbergen, J., Komdeur, J., Kats, R. & Berg, M. 2001. Egg discrimination in the Australian reed warbler (Acrocephalus australis): rejection response toward model and conspecific eggs depending on timing and mode of artificial parasitism. Behavioral Ecology, 12, 8e15. Zar, J. H. 1996. Biostatistical Analysis. 3rd edn. Upper Saddle River, New Jersey: Prentice Hall.
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