Cross‐fostering reveals sources of variation in innate immunity and hematocrit in nestling tree swallows Tachycineta bicolor

Innate immunity plays an important role in protecting birds early in development. The strength of innate immunity in nestlings appears to be affected by heritable and early maternal effects, as well as later environmental effects. To explore factors affecting innate immunity, we conducted a partial cross‐fostering experiment in nestling tree swallows Tachycineta bicolor. The ability of nestling blood plasma to kill Escherichia coli in vitro was influenced by heritable or early maternal effects and was unrelated to body condition of nestlings or brood size of the foster nest. The relative importance of heritable and early maternal effects versus environmental effects on nestling hematocrit (packed volume of red blood cells) and body condition were also examined. Hematocrit was significantly influenced by the nest in which nestlings were reared, though it was not related to nestling body condition. Body condition was significantly influenced by the nest in which nestlings were reared, though it was also weakly influenced by heritable or early maternal effects. In summary, heritable and early maternal effects are stronger predictors than rearing conditions of innate immunity in nestling tree swallows. Furthermore, hematocrit is largely environmentally determined but is not related to body condition or innate immunity, as measured in this study.     

Offspring quality and tick infestation load in brood rearing great tits Parus major

Although the impact of nest-dwelling ectoparasites on nestlings in altricial birds is relatively well documented, little information is available on the fitness consequences of bird–ectoparasite systems with limited ectoparasite transmission between parents and their offspring. In this particular context we tested the hypothesis that parental infestation by a haematophageous field ectoparasite, the sheep tick Ixodes ricinus , in a hole-breeding passerine bird, the great tit Parus major , ultimately leads to the reduction of nestling quality. Observational data of a long-term study show that 15 days after hatching, nestlings' body condition (body mass corrected for body size) was negatively associated with the tick load of the mother, while no association was found with tick load of the father. A field experiment was conducted to test if adult infestation by ticks leads to a reduction in offspring quality. The field experiment consisted of a two by two factorial design: neither, one or both parents were infested with a high tick load (12 nymphs) when nestlings were eight days old. Four days after manipulation of the parents, the effect of the treatment on nestlings' health was assessed by measuring haematological and biochemical parameters. Body condition of nestlings was monitored from three days after hatching, until seven days after tick manipulation of the parents. Although parental tick manipulation resulted in successful tick feeding, nestlings' health parameters were not affected. We therefore suggest that the negative association between nestling condition and parental tick load does not reflect a causal effect of parasites, but either reflects a common environmental factor affecting parental infestation levels and offspring condition, or reflects parental quality. We propose different explanations why this association is expressed in female parents only.     

Differential virulence in two congeneric ticks infesting songbird nestlings

Virulence is part of the proximate base to the understanding of host-parasite interactions, and therefore it is crucial to interpret parasite-induced damage as an outcome of co-evolution between parasite and host. We experimentally investigated the impact of 2 congeneric ticks with contrasting transmission modes and host specificity, on the health of songbird nestlings (Parus major). The nidicolous Ixodes arboricola lives in tree holes where it obtains blood from adult birds and their nestlings, hence is prone to vertical transmission. All developmental stages of the field-dwelling Ixodes ricinus feed on different host taxa, hence are prone to horizontal transmission. Within each nest we infested 3 nestlings with varying loads of I. ricinus and 3 with Ixodes arboricola. Ixodes arboricola had no negative effects on nestling health (haematocrit, inflammation, body size, body condition). Body size was positively associated with the number of feeding I. arboricola adults. In contrast, I. ricinus infestations by nymphs and adult ticks led to haematocrit reductions. Furthermore, I. arboricola was a more prudent feeder, with longer feeding durations for smaller bloodmeals. Although the negative effects on nestling health were limited, our results are in line with current theories, stating that virulence increases with horizontal transmission, reduced dependence on individual hosts and reduced relatedness among co-exploiting ticks.     

Behavioural responses to ectoparasites in pied flycatchers Ficedula hypoleuca: an experimental study

Nests of cavity‐nesting birds usually harbor some species of haematophagous ectoparasites that feed on the incubating adults and nestlings. Given the negative impact of ectoparasites on nestlings there will be selection on hosts to reduce parasite infestations through behavioural means. We have experimentally reduced the abundance of all ectoparasites in nests of pied flycatchers Ficedula hypoleuca to explore both whether there are changes in the frequency and duration of putative anti‐parasite behaviours by tending adults, as well as whether such anti‐parasite behaviours are able to compensate for the deleterious effects that parasites may have on nestlings. Heat treatment of nests substantially decreased the density of ectoparasites, and thereby positively affected nestling growth. The frequency and intensity of female grooming and nest sanitation behaviours during the incubation and nestling periods decreased as a consequence of the experimental reduction of ectoparasite infestation. Although nestlings begged more intensely in infested nests, the experiment had no significant effect on parental provisioning effort. Reduction of parasites resulted in larger nestlings shortly before fledging and increased fledging success. This study shows a clear effect of a complete natural nest ectoparasite fauna on parental behaviour at the nest and nestling growth in a cavity‐nesting bird. Although ectoparasites induce anti‐parasite behaviours in females, these behaviours are not able to fully remove parasite's deleterious effects on nestling growth and survival.     

Experimental study of micro-habitat selection by ixodid ticks feeding on avian hosts

Mechanisms of on-host habitat selection of parasites are important to the understanding of host-parasite interactions and evolution. To this end, it is important to separate the factors driving parasite micro-habitat selection from those resulting from host anti-parasite behaviour. We experimentally investigated whether tick infestation patterns on songbirds are the result of an active choice by the ticks themselves, or the outcome of songbird grooming behaviour. Attachment patterns of three ixodid tick species with different ecologies and host specificities were studied on avian hosts. Ixodes arboricola, Ixodes ricinus and Ixodes frontalis were put on the head, belly and back of adult great tits (Parus major) and adult domestic canaries (Serinus canaria domestica) which were either restricted or not in their grooming capabilities. Without exception, ticks were eventually found on a bird’s head. When we gave ticks full opportunities to attach on other body parts – in the absence of host grooming – they showed lower attachment success. Moreover, ticks moved from these other body parts to the host's head when given the opportunity. This study provides evidence that the commonly observed pattern of ticks feeding on songbirds’ heads is the result of an adaptive behavioural strategy. Experimental data on a novel host species, the domestic canary, and a consistent number of published field observations, strongly support this hypothesis. We address some proximate and ultimate causes that may explain parasite preference for this body part in songbirds. The link found between parasite micro-habitat preference and host anti-parasite behaviour provides further insight into the mechanisms driving ectoparasite aggregation, which is important for the population dynamics of hosts, ectoparasites and the micro-pathogens for which they are vectors.     

Habitat-specific effects of a food supplementation experiment on immunocompetence in Eurasian Magpie Pica pica nestlings

Parasite pressure and nutrition are two of the most important factors affecting the trade-off between nestling growth and immune development. During development, energy and nutrients are often limited, and nestlings should only dedicate differentially more valuable resources to their immune system when the associated benefits are high (i.e. in situations of an increased risk of parasitism). In this study, we manipulated nutritional condition of Eurasian Magpie Pica pica nestlings by providing a food supplement. Additionally, the study area was subdivided into two categories of habitat (irrigated and arid) based on the presence of irrigation canals. Nestling diet composition was more varied in the irrigated compared with in the arid habitat. In addition, nestlings of the irrigated habitat showed a significantly higher infestation of both ectoparasites and blood parasites and a significantly higher cell-mediated immunity, but lower tarsus length compared with nestlings of the arid habitat. Food supplementation to nestlings did not affect tarsus length, but increased nestling cell-mediated immunity in the arid habitat only. Based on the recent demonstrated trade-off between growth and immunocompetence in nestlings, we suggest that differences between habitats in nestling diet and parasite prevalence may have caused different priority rules in the allocation of resources between both fitness traits.     

Field experiments on the effect of ticks on breeding success and chick health of cattle egrets

Abstract Two experiments compared broods that were naturally tick-infested with an equal number that were rendered tick-free by application of an acaricide to their nests and of barriers against further infestation. In the first experiment conducted in 1991–92 nestlings in tick-infested broods had up to 159 larval ticks at once and a mean infestation of 23.6 larval ticks per chick per day. The chick's mean tick load was inversely correlated with its longevity. Chicks that survived to at least 18 days post-hatching had significantly lower larval tick loads than those that died by 18 days, excluding the third-hatched chick in each brood whose survival rate was low irrespective of tick-infestation. At 7 days post-hatching, tick-infested chicks had a lower haematocrit and higher polychromasia than tick-free chicks. I infer that blood loss anaemia caused the deaths of tick-infested chicks in their first week. None died in their second week and the demise of those in their third week may have been due to the paralysis manifested prior to their death. I conclude that heavy tick infestation of their chicks reduced the breeding success of the parent egrets below that of the egrets whose chicks were kept free of ticks. In the second experiment in the 1992–93 season the mean level of tick infestation was much lower (5.1 per chick) and these chicks survived equally with tick-free chicks through fledging.     

Immunocompetence of nestling great tits in relation to rearing environment and parentage

Theoretical models of host–parasite coevolution assume a partially genetic basis to the variability in susceptibility to parasites among hosts, for instance as a result of genetic variation in immune function. However, few empirical data exist for free-living vertebrate hosts to support this presumption. In a cross-fostering experiment with nestling great tits, by comparing nestlings of the same origin we investigated (i) the variance in host resistance against an ectoparasite due to a common genetic origin, (ii) the effect of ectoparasite infestation on cell-mediated immunity and (iii) the variance in cell-mediated immunity due to a common genetic origin. Ectoparasitic hen fleas can impair the growth of nestling great tits and nestling growth was therefore taken as a measure of host susceptibility. A common origin did not account for a significant part of the variation in host susceptibility to fleas. There was no significant overall effect of fleas on nestling growth or cell-mediated immunity, as assessed by a cutaneous hypersensitivity response. A common rearing environment explained a significant part of the variation in cell-mediated immunity among nestlings, mainly through its effect on nestling body mass. The variation in cell-mediated immunity was also related to a common origin. However, the origin-related variation in body mass did not account for the origin-related differences in cell-mediated immunity. The results of the present study thus suggest heritable variation in cell-mediated immunity among nestling great tits.     

Tick saliva in anti-tick immunity and pathogen transmission

When feeding on vertebrate host ticks (ectoparasitic arthropods and potential vectors of bacterial, rickettsial, protozoal, and viral diseases) induce both innate and specific acquired host-immune reactions as part of anti-tick defenses. In a resistant host immune defense can lead to reduced tick viability, sometimes resulting in tick death. Tick responds to the host immune attack by secreting saliva containing pharmacologically active molecules and modulating host immune response. Tick saliva-effected immunomodulation at the attachment site facilitates both tick feeding and enhances the success of transmission of pathogens from tick into the host. On the other hand, host immunization with antigens from tick saliva can induce anti-tick resistance and is seen to be able to induce immunity against pathogens transmitted by ticks. Many pharmacological properties of saliva described in ticks are shared widely among other blood-feeding arthropods.     

An experimental test to compare potential and realised specificity in ticks with different ecologies

The majority of studies on ecological specialisation rely on data reflecting realised specificity, without considering species’ potential specificity. Most species of ticks, a large family of hematophagous ectoparasites, have a narrow host range in nature, but it is unclear whether this is due to host-driven adaptations or other processes (such as off-host abiotic environment). We investigated the potential specificity of two tick species with contrasting ecology by infesting three avian host species that occur in the same off-host macrohabitat but are unequally infested by the ticks in nature (i.e. have contrasting realised specificity). The endophilic specialist tick Ixodes arboricola resides inside the hosts’ nest and has high realised host specificity, whereas the exophilic generalist tick I. ricinus encounters hosts in the field and has very low realised specificity. As hosts, we used great tits (frequently infested by both tick species), blackbirds (frequently infested by I. ricinus but never by I. arboricola) and great spotted woodpeckers (no ticks of either species have been reported). If realised specificity is constrained by host-driven adaptations there should be no differences between potential and realised specificity, whereas if realised specificity is constrained by other processes potential specificity and realised specificity should be different. We found that attachment rates and weight during feeding of I. arboricola were lower on blackbirds than on great tits, whereas there were no such differences for I. ricinus. No ticks of either species attached to woodpeckers. These results indicate that realised host specificity of ticks is, at least partially, constrained by host-driven adaptations. This specificity therefore strongly depends on the ticks’ encounter rates with particular host types, which are affected by the ticks’ off-host ecological requirements, behaviour and life-history characteristics.     

Adaptive host-abandonment of ectoparasites before fledging? Within-brood distribution of nest mites in house sparrow broods

We studied the within-brood distribution of a haematophagous mite Pellonyssus reedi living on nestling house sparrows (Passer domesticus) near the time of fledging. We measured the natural level of infestation of individual nestlings, and determined the feeding efficiency of mites, by scoring their feeding status. Within-brood distribution of mite loads was unrelated to nestling body mass, tarsus length, or immunocompetence. These results did not support parasite preference for large or susceptible hosts. Mite feeding-efficiency was also unrelated to these nestling characteristics, confirming that large nestlings or nestlings with less-developed immunocompetence did not provide superior feeding conditions for mites. Therefore, our results do not support the hypothesis that within-brood distribution of avian ectoparasites is explained by the parasites' preferences for characteristics, such as large body size or low immunocompetence, that make nestlings suitable hosts. On the other hand, we found that mite loads were negatively correlated with nestling age and feather length, suggesting that nestlings closer to fledging harbored fewer mites then their less-developed nestmates. Furthermore, feather length had a stronger relationship with parasite distribution than did nestling age. We presume, therefore, that feather characteristics, i.e., length, may serve as a signal for mites to perceive the ready-to-fledge state of nestlings, inducing abandonment behavior. These results support another, largely neglected hypothesis, i.e., that the avoidance or abandonment of those nestlings that are close to fledging may also explain the parasites' distribution in a brood. This hypothesis is based on the argument that many nest-dwelling ectoparasites breed in the nest material and emerge only periodically to feed on nestlings. In such parasites, the ability to recognize and avoid mature fledglings can be adaptive because this may help the parasites to avoid their removal from the nest so they can continue to reproduce by feeding on unfledged chicks of the current or later broods. Our results suggest that adaptive host-abandonment by nest-dwelling ectoparasites can influence within-brood parasite distributions around the time of fledging.     

Cost of parasitism and host immune defence in the sand martin Riparia riparia: a role for parent-offspring conflict?

Parent-offspring conflict may arise because the lifetime reproductive success of the parent is more influenced by its life span than by reproductive success during a particular reproductive event, while the fitness of an offspring depends firstly on its own survival as a juvenile and only subsequently on its own reproductive success. The naive immune system of young animals may allow offspring to be much more affected by parasites than their parents, and thus cause an initial asymmetry in a potential parent-offspring conflict. We investigated this type of conflict by assessing the health status and the immune response of parent and offspring sand martins Riparia riparia infested with manipulated loads of ticks Ixodes lividus (nests either treated with pyrethrin, water, or just visited). The prevalence and the intensity of tick infestations differed among treatments, with low tick loads in nests with the pyrethrin treatment. Ticks reduced the reproductive success of the host and increased offspring wing length. Broods with ticks had higher leukocyte concentrations and concentrations of immunoglobulins. The concentration of immunoglobulins in nestlings was negatively related to brood size and nestling tarsus length. Nestlings receiving the control treatments had a positive association between wing length and the concentration of immunoglobulins and a negative association between tarsus length and immunoglobulins. In contrast, adult sand martins did not respond to the parasite treatment in terms of immune response. Hence, the naive immune system of nestlings may be the crucial factor causing the parent-offspring conflict over costs of parasitism to be resolved to the advantage of parents that may sacrifice nestlings in heavily parasitized nests. Received: 30 March 1998 / Accepted: 5 December 1998     

Do leucocytes reflect condition in nestling burrowing parrots Cyanoliseus patagonus in the wild?

The different leucocyte types are an important part of the immune system. Thus, they have been used in ecological studies to assess immune function and physiological stress in wild birds. It is generally assumed that increased stress and decreased condition are associated with an increase in the ratio of heterophils to lymphocytes, the H/L ratio. We studied leucocyte profiles in relation to body condition in nestling Burrowing Parrots (Cyanoliseus patagonus) in North-eastern Patagonia, Argentina. As in other wild parrots, heterophils were the most numerous leucocyte type, suggesting strong investment into innate immunity. Leucocyte profiles did not change with the age, while nestlings in better body condition increased the number of heterophils. Because the number of lymphocytes was independent of body condition, as a result we observed a positive correlation between body condition and the H/L ratio. The total number of leucocytes relative to erythrocytes increased in nestlings in better body condition, indicating a larger overall investment into immune function in well-nourished nestlings. The observed heterophilic profiles of nestling Burrowing Parrots together with the positive relationship between H/L ratio and body condition may indicate a favoured investment in a robust innate immunity that reduces the risk of infection taking hold in these long-lived birds.     

Experimental manipulation of temperature reduce ectoparasites in nests of blue tits Cyanistes caeruleus

Several models predict changes in the distributions and incidences of diseases associated with climate change. However, studies that investigate how microclimatic changes may affect host–parasite relationships are scarce. Here, we experimentally increased the temperature in blue tit Cyanistes caeruleus nest boxes during their breeding season to determine its effects on the parasitic abundance (i.e. of nest‐dwelling ectoparasites, blood‐sucking flying insects and hemoparasites) in nests and the host condition of nestlings and adults. The temperature was increased using heat mats placed underneath the nest material, which resulted in an average temperature increase of 3ºC and a reduction in relative humidity of about six units. The abundance of mites Dermanyssus gallinoides and blowfly pupae Protocalliphora azurea was significantly reduced in heated nest boxes. Although not statistically significant, a lower prevalence of flea larvae Ceratophyllus gallinae was also found in heated nests. However, heat treatment did not affect hemoparasite infection of adult blue tits or the body condition of adult and nestling blue tits. In conclusion, heat treatment in blue tit nests reduced nest‐dwelling ectoparasites yet without any apparent benefit for the host.     

Cost of reproduction, T-lymphocyte mediated immunocompetence and health status in female and nestling barn swallows Hirundo rustica

We investigate the trade-off between reproductive effort, health status and T-lymphocyte acquired immunity in female and nestling barn swallows Hirundo rustica using a brood size manipulation experiment. Maternal and total feeding effort increased with experimental brood size. Parents did not fully compensate for the increased food demand of the enlarged broods and as a consequence the per capita feeding rate of nestlings decreased with increasing experimental brood size. Body mass and a measure of T-cell mediated immunity in 12 days old nestlings also decreased with increasing experimental brood size. Different leucocyte concentrations and the heterophile/lymphocyte ratio – an index of stress – of nestlings did not change in relation to experimental brood size, suggesting that within brood competition did not affect stress to nestlings. The brood size manipulation had a significant effect on maternal T-cell mediated immunity, measured by the phytohemagglutinin skin test, but not on maternal body mass, haematocrit or differential or total white blood cell counts. Our results seem to support the prediction that under mild work stress females respond first by reducing the energetically expensive acquired immunity. Different leucocyte types and the heterophile/lymphocyte ratio appear less sensitive to parental workload.     

Early maternal effects and antibacterial immune factors in the eggs,nestlings and adults of the barn swallow

Transfer of immune factors via the egg may represent a maternal adaptation enhancing offspring survival. Lysozyme is a major component of maternal antibacterial immunity which is transferred to the eggs in birds. In a population of barn swallows (Hirundo rustica), lysozyme activity declined during the prelaying and laying periods in females but not in males. Egg hatching failure decreased with maternal lysozyme activity. The first eggs in a clutch contained more lysozyme and produced nestlings with larger lysozyme activity when 5 days old than last‐laid ones. In a cross‐fostering experiment where brood size was manipulated, nestling origin but not post‐manipulation brood size affected lysozyme activity. Hence, maternal lysozyme varies during the breeding season and may differentially enhance antibacterial immune defence of the eggs and nestlings in relation to laying order. These findings suggest that offspring innate immunity is influenced by early maternal effects.     

Ticks (Acari: Ixodoidea) in China: Geographical distribution,host diversity,and specificity

Ticks are obligate blood‐sucking ectoparasites, which not only directly damage through bites but also transmit many pathogens. China has a high diversity of tick species, 125 species have been reported, including 111 hard tick and 14 soft tick species. Many of the ticks are important vectors of pathogens, resulting in zoonoses. The dynamics of ticks are affected by both the host and habitat environment. However, systematic studies on the geographical distribution, host diversity, and specificity of ticks are limited in China. To achieve this goal, the relevant available data were summarized and analyzed in this study. Ticks are distributed in all parts of China and Xinjiang has the most records of ticks. The distribution of ticks in adjacent areas is similar, indicating that the habitat environment affects their distribution. Most ticks are widely distributed, whereas some species are endemic to their distributed regions. Ticks are parasitic on mammals, birds, and reptiles, of which mammals are the main host species. Overall, most ticks parasitize different hosts, only a few ticks have strict host specificity, such as ticks that are specifically parasitic on reptiles and bats. In addition, environmental changes and control efforts also influence the dynamics of ticks. These results can better reveal tick biological traits and are valuable for tick control.     

Hatching asynchrony and brood reduction influence immune response in Common Kestrel Falco tinnunculus nestlings

The onset of incubation before the end of laying imposes asynchrony at hatching and, therefore, a size hierarchy in the brood. It has been argued that hatching asynchrony might be a strategy to improve reproductive output in terms of quality or quantity of offspring. However, little is known about the mediating effect of hatching asynchrony on offspring quality when brood reduction occurs. Here, we investigate the relationship between phenotypic quality and hatching asynchrony in Common Kestrel Falco tinnunculus nestlings in Spain. Hatching asynchrony did not increase breeding success or nestling quality. Furthermore, hatching asynchrony and brood reduction had different effects on nestlings’ phytohaematogglutinin (PHA)‐mediated immune response and nestling growth. In asynchronous and reduced broods (in which at least one nestling died), nestlings showed a stronger PHA‐mediated immune response and tended to have a smaller body size compared with nestlings raised in synchronous and reduced broods. When brood reduction occurred in broods hatched synchronously, there was no effect on nestling size, but nestlings had a relatively poor PHA‐mediated immune response compared with nestlings raised in asynchronous and reduced broods. We suggest that resources for growth can be directed to immune function only in asynchronously hatched broods, resulting in improved nestling quality, as suggested by their immune response. We also found that males produced a greater PHA‐mediated immune response than females only in brood‐reduced nests without any effect on nestling size or condition, suggesting that females may trade off immune activities and body condition, size or weight. Overall, our results suggest that hatching pattern and brood reduction may mediate resource allocation to different fitness traits. They also highlight that the resolution of immune‐related trade‐offs when brood reduction occurs may differ between male and female nestlings.     

Ectoparasitism and the role of green nesting material in the European starling

Summary The use of green nesting material is widespred among birds. Recent evidence suggests that birds use secondary chemicals contained in green plants to control ectoparasites. We manipulated green nesting material and ectoparasites of European starlings (Sturnus vulgaris) to test two hypotheses: (1) ectoparasites adversely affect prefledging survival and morphometrics or postfledging survival, and (2) green nesting material ameliorates the effects of ectoparasites. We recorded fat score, numbers of scabs, tarsal length, body mass, and hematocrit level on each nestling 17 days after hatching. We also fitted each nestling with unique patagial tags and resighted the starlings for 6–8 weeks after fledging to estimate survival and sighting rates. Nests devoid of green nesting material and dusted with the insecticide, carbaryl, had fewer high ectoparasite infestations, and nestlings had significantly lower scab scores, and significantly higher body masses than nestlings in undusted boxes. However, there was no difference in postfledging survival between birds from carbaryl-treated and undusted nests. There also was no difference in prefledging survival and morphometrics or postfledging survival between nestlings from boxes with and without green nesting material. These results do not support the hypothesis that starlings use green nesting material to control nest ectoparasites. We suggest an alternative hypothesis; green nesting material is used for mate selection or pairbonding in the starling.     

Phenotypic variation in nestlings of a bird of prey under contrasting breeding and diet conditions

Environmental conditions often vary in space and time, and this may explain variation in the expression of phenotypic traits related to individual quality, such as ornamental coloration. Furthermore, the direction and strength of the relationship between coloured trait expression and individual quality might vary under contrasting conditions. These issues have been explored in adult birds but much less so in nestlings, which are more likely to experience different selective pressures and different physiological trade‐offs than adults. Here, we empirically investigated the effects of contrasting breeding and diet conditions on the expression of carotenoid‐based colour traits displayed by marsh harrier (Circus aeruginosus) nestlings. We studied the variation in coloration, body condition, and immune responsiveness of nestlings in four populations over a 5‐year period. We characterized spatiotemporal differences in rearing conditions experienced by C. aeruginosus nestlings in terms of breeding (laying date, clutch size, and number of nestlings hatched and fledged) and diet (percentage of mammal in diet and prey diversity) conditions. We found that breeding conditions influenced the co‐variation between coloration and immune responsiveness in female nestlings, and that diet conditions influenced the condition‐dependence of nestling coloration in later‐hatched nestlings. In addition, breeding conditions influenced nestling body condition and immune responsiveness, whereas diet conditions influenced nestling coloration and body condition. Our study highlights that nestling phenotype (levels of signalling, circulating carotenoids, and immunity) varies both spatially and temporally, and that some of this variation is related to differences in breeding and diet conditions. Moreover, under contrasting conditions, the direction of the relationships between nestling carotenoid‐based coloration and nestling quality may also vary. In order to fully understand the evolution and maintenance of colour traits in nestling birds, studies and experiments should ideally be replicated under contrasting rearing conditions. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, ?? , ??–??.