Mercury Uptake Affects the Development of Larus fuscus Chicks.

Current emission and mobilization rates of mercury (Hg) in the environment pose extensive threats to both wildlife and human health. Assessing the exposure risk and effects of Hg contamination in model species such as seabirds is essential to understand Hg risks at the population and ecosystem levels. The lesser black-backed gull (Larus fuscus), a generalist seabird species, is an excellent model species because it forages in both marine and terrestrial habitats, which in turn differ in their Hg exposure risk. To identify possible deleterious effects of Hg exposure on developing L. fuscus chicks, a dietary experiment was carried out and chicks were provided a marine, terrestrial, or mixed diet. The effects of embryonic and dietary Hg exposure on chick body condition and physiological state were assessed at different developmental stages until fledging age (30 d). Overall physiological condition was lower in chicks fed a predominantly marine diet, which coincided with higher Hg loads in blood and primary feathers. However, no effect of dietary uptake of Hg was observed on body condition or in terms of genotoxic damage. Body condition and genotoxic damage correlated instead with Hg exposure during embryonic development, which seems to indicate that embryonic exposure to Hg may result in carry-over effects on later chick development. Environ Toxicol Chem 2020;39:2008-2017. © 2020 SETAC.

Diet contributes to urban-induced alterations in gut microbiota: experimental evidence from a wild passerine.

Urban sprawl increasingly affects the ecology of natural populations, including host-microbiota interactions, with observed differences in the gut microbiota between urban and rural hosts. While different mechanisms could explain this pattern, dietary uptake constitutes a likely candidate. To assess the contribution of diet in explaining urban-rural variation in gut microbiota, we performed an aviary experiment in which urban and rural house sparrows were fed with mimics of urban or rural diets. Before the experiment, rural sparrows hosted more diverse gut communities, with a higher relative abundance of Enterococcaceae and Staphylococcaceae and lower abundance of genes involved in xenobiotic degradation and lipid metabolism than their urban counterparts. The experimental diets significantly altered gut microbiota α- and ß-diversity and taxonomic composition, with the strongest shifts occurring in individuals exposed to contrasting diets. Overall, diet-induced shifts resembled initial differences between free-ranging urban and rural hosts. Furthermore, rural diet had a positive impact on urban host body mass but only in hosts with the highest initial gut diversity. Overall, our results indicate that diet constitutes an important factor contributing to differences in gut microbiota along the urbanization gradient and provide new insights on possible fitness consequences of a reduced gut diversity in urban settings.

Assay optimisation and age-related baseline variation in biochemical markers in Lesser Black-backed gulls.

Free-ranging animals are often used as bioindicators of both short- and long-term changes in ecosystem health, mainly to detect the presence and effects of contaminants. Birds, and gulls in particular, have been used as bioindicators over a broad range of marine and terrestrial ecosystems. In this study, we standardise the conditions for the use of a suite of biochemical markers in non-destructive matrices of Lesser Black-backed Gull (Larus fuscus) to facilitate future biomonitoring of marine and terrestrial contaminants. We characterized cholinesterase (ChE) in plasma and optimized assay conditions for ChE activity as a marker of neurotoxic damage. Moreover, we quantified variation in activity of ChE, lactate dehydrogenase (LDH), glutathione-S-transferase (GST) and catalase (CAT) as well as variation ranges of lipid peroxidation (LPO), in free-ranging adults and captive chicks. The main ChE form present in plasma of both adults and chicks was butyrylcholinesterase (BChE) followed by acetylcholinesterase (AChE), whose relative proportion in plasma tended to decrease with increased chick age. LPO levels and GST activity in blood cells (BCs) decreased significantly with increasing chick age, while BChE and LDH activity in plasma were not age-dependent. CAT in BCs tended to decline non-significantly in older chicks. Results of this study underscore the importance of standardising assay conditions and assessing intrinsic baseline variation in biochemical markers, before biochemical quantification. Data presented here provide a foundation for future use of BChE and LDH activity in plasma, as well as oxidative stress markers (LPO, CAT and GST) in BCs, to monitor environmental stress effects in Lesser Black-backed gulls.

Do wild-caught urban house sparrows show desensitized stress responses to a novel stressor?

While urbanization exposes individuals to novel challenges, urban areas may also constitute stable environments in which seasonal fluctuations are buffered. Baseline and stress-induced plasma corticosterone (cort) levels are often found to be similar in urban and rural populations. Here we aimed to disentangle two possible mechanisms underlying such pattern: (i) urban environments are no more stressful or urban birds have a better ability to habituate to stressors; or (ii) urban birds developed desensitized stress responses. We exposed wild-caught urban and rural house sparrows (Passer domesticus) to combined captivity and diet treatments (urban versus rural diet) and measured corticosterone levels both in natural tail feathers and in regrown homologous ones (cortf). Urban and rural house sparrows showed similar cortf levels in the wild and in response to novel stressors caused by the experiment, supporting the growing notion that urban environments are no more stressful during the non-breeding season than are rural ones. Still, juveniles and males originating from urban populations showed the highest cortf levels in regrown feathers. We did not find evidence that cortf was consistent within individuals across moults. Our study stresses the need for incorporating both intrinsic and environmental factors for the interpretation of variation in cortf between populations.

Supplementary feeding increases nestling feather corticosterone early in the breeding season in house sparrows.

Several studies on birds have proposed that a lack of invertebrate prey in urbanized areas could be the main cause for generally lower levels of breeding success compared to rural habitats. Previous work on house sparrows Passer domesticus found that supplemental feeding in urbanized areas increased breeding success but did not contribute to population growth. Here, we hypothesize that supplementary feeding allows house sparrows to achieve higher breeding success but at the cost of lower nestling quality. As abundant food supplies may permit both high- and low-quality nestlings to survive, we also predict that within-brood variation in proxies of nestling quality would be larger for supplemental food broods than for unfed broods. As proxies of nestling quality, we considered feather corticosterone (CORT f), body condition (scaled mass index, SMI), and tarsus-based fluctuating asymmetry (FA). Our hypothesis was only partially supported as we did not find an overall effect of food supplementation on FA or SMI. Rather, food supplementation affected nestling phenotype only early in the breeding season in terms of elevated CORT f levels and a tendency for more variable within-brood CORT f and FA. Early food supplemented nests therefore seemed to include at least some nestlings that faced increased stressors during development, possibly due to harsher environmental (e.g., related to food and temperature) conditions early in the breeding season that would increase sibling competition, especially in larger broods. The fact that CORT f was positively, rather than inversely, related to nestling SMI further suggests that factors influencing CORT f and SMI are likely operating over different periods or, alternatively, that nestlings in good nutritional condition also invest in high-quality feathers.

Offspring Hg exposure relates to parental feeding strategies in a generalist bird with strong individual foraging specialization.

Generalist species can potentially exploit a wide variety of resources, but at the individual level they often show a certain degree of foraging specialization. Specific foraging strategies, however, may increase exposure to environmental contaminants that can alter the cost-benefit balance of consuming particular food items. The Lesser Black-backed Gull (Larus fuscus) is known to opportunistically feed on a wide range of marine and terrestrial prey that differ in contaminant load, such as mercury (Hg) that strongly biomagnifies through the aquatic food web. The hypothesis tested in this study were: i) a predominant use of marine prey by females during egg-formation and by both parents during chick rearing increases the exposure to Hg during embryonic development and chick growth, and ii) this affects parental investment in clutch volume, chick growth and body condition. Total Hg burden and isotopic signatures of carbon (δ13C) and nitrogen (δ15N) were determined for eggs, down feathers, and primary feathers of L. fuscus chicks collected at a coastal colony in Belgium. As expected, eggs and feathers of chicks from parents with a stable isotope signature that suggested a predominantly marine diet had higher levels of Hg. The use of marine resources by females during the egg-formation period positively correlated to maternal investment in egg size, though entailing the cost of increased Hg-concentrations which in turn negatively affected clutch volume. Furthermore, it is shown that the use of chick down feathers is a suitable matrix to non-lethally estimate Hg concentrations in eggs. Contrary to our expectations, no relationship between Hg exposure and chick growth or chick body condition was found, which may be due the low concentrations found. We conclude that currently Hg contamination does not constitute a risk for development and condition of L. fuscus offspring at the levels currently observed at the Belgian coast.

Cooperative breeding shapes post-fledging survival in an Afrotropical forest bird.

For avian group living to be evolutionary stable, multiple fitness benefits are expected. Yet, the difficulty of tracking fledglings, and thus estimating their survival rates, limits our knowledge on how such benefits may manifest postfledging. We radio-tagged breeding females of the Afrotropical cooperatively breeding Placid greenbul (Phyllastrephus placidus) during nesting. Tracking these females after fledging permitted us to locate juvenile birds, their parents, and any helpers present and to build individual fledgling resighting datasets without incurring mortality costs or causing premature fledging due to handling or transmitter effects. A Bayesian framework was used to infer age-specific mortality rates in relation to group size, fledging date, maternal condition, and nestling condition. Postfledging survival was positively related to group size, with fledglings raised in groups with four helpers showing nearly 30% higher survival until independence compared with pair-only offspring, independent of fledging date, maternal condition or nestling condition. Our results demonstrate the importance of studying the early dependency period just after fledging when assessing presumed benefits of cooperative breeding. While studying small, mobile organisms after they leave the nest remains highly challenging, we argue that the telemetric approach proposed here may be a broadly applicable method to obtain unbiased estimates of postfledging survival.

Great spotted cuckoo nestlings have no antipredatory effect on magpie or carrion crow host nests in southern Spain.

Host defences against cuckoo parasitism and cuckoo trickeries to overcome them are a classic example of antagonistic coevolution. Recently it has been reported that this relationship may turn to be mutualistic in the case of the carrion crow (Corvus corone) and its brood parasite, the great spotted cuckoo (Clamator glandarius), given that experimentally and naturally parasitized nests were depredated at a lower rate than non-parasitized nests. This result was interpreted as a consequence of the antipredatory properties of a fetid cloacal secretion produced by cuckoo nestlings, which presumably deters predators from parasitized host nests. This potential defensive mechanism would therefore explain the detected higher fledgling success of parasitized nests during breeding seasons with high predation risk. Here, in a different study population, we explored the expected benefits in terms of reduced nest predation in naturally and experimentally parasitized nests of two different host species, carrion crows and magpies (Pica pica). During the incubation phase non-parasitized nests were depredated more frequently than parasitized nests. However, during the nestling phase, parasitized nests were not depredated at a lower rate than non-parasitized nests, neither in magpie nor in carrion crow nests, and experimental translocation of great spotted cuckoo hatchlings did not reveal causal effects between parasitism state and predation rate of host nests. Therefore, our results do not fit expectations and, thus, do not support the fascinating possibility that great spotted cuckoo nestlings could have an antipredatory effect for host nestlings, at least in our study area. We also discuss different possibilities that may conciliate these with previous results, but also several alternative explanations, including the lack of generalizability of the previously documented mutualistic association.

Use of LC-MS-MS as an alternative to currently available immunoassay methods to quantitate corticosterone in egg yolk and albumen.

Corticosterone (CORT) is the dominant plasma glucocorticoid in birds. There has been increasing interest in the function of CORT in avian egg yolk and in the potential to use CORT concentrations in eggs to quantify stress and to assess the effect of maternal stress on offspring. The concentration of CORT in egg yolk is most frequently assessed using enzyme or radioimmunoassays, alone or in combination with high-performance liquid chromatography. However, the quantification of CORT is frequently hampered by the presence of high concentrations of other steroid hormones which cross-react with the CORT antibody. As an alternative, we developed a sensitive and specific LC-MS-MS method. The sample-preparation procedure consisted of a protein-lipid precipitation step, followed by defatting and clean-up using a C18 SPE column. Chromatography was performed on an Acquity C18 BEH column (50 mm × 2.1 mm i.d., dp: 1.7 µm, run-time: 6 min), using 0.1% formic acid in both water (A) and acetonitrile (B) as mobile phases. The MS-MS instrument was operated in the positive-electrospray-ionization mode. The method was validated in-house according to European Guidelines (linearity, accuracy and precision, limits of quantification and detection, specificity, stability) and the results fell within the accepted ranges. The method was successfully used for the analysis of CORT in yolk and albumen of eggs collected from eight breeding lesser black-backed gulls at a Flemish coastal colony. CORT concentrations were in the range 42.4-166.3 pg g(-1) in albumen and < LOQ (75 pg g(-1))-762.5 pg g(-1) in yolk.

Great spotted cuckoo fledglings often receive feedings from other magpie adults than their foster parents: which magpies accept to feed foreign cuckoo fledglings?

Natural selection penalizes individuals that provide costly parental care to non-relatives. However, feedings to brood-parasitic fledglings by individuals other than their foster parents, although anecdotic, have been commonly observed, also in the great spotted cuckoo (Clamator glandarius)--magpie (Pica pica) system, but this behaviour has never been studied in depth. In a first experiment, we here show that great spotted cuckoo fledglings that were translocated to a distant territory managed to survive. This implies that obtaining food from foreign magpies is a frequent and efficient strategy used by great spotted cuckoo fledglings. A second experiment, in which we presented a stuffed-cuckoo fledgling in magpie territories, showed that adult magpies caring for magpie fledglings responded aggressively in most of the trials and never tried to feed the stuffed cuckoo, whereas magpies that were caring for cuckoo fledglings reacted rarely with aggressive behavior and were sometimes disposed to feed the stuffed cuckoo. In a third experiment we observed feedings to post-fledgling cuckoos by marked adult magpies belonging to four different possibilities with respect to breeding status (i.e. composition of the brood: only cuckoos, only magpies, mixed, or failed breeding attempt). All non-parental feeding events to cuckoos were provided by magpies that were caring only for cuckoo fledglings. These results strongly support the conclusion that cuckoo fledglings that abandon their foster parents get fed by other adult magpies that are currently caring for other cuckoo fledglings. These findings are crucial to understand the co-evolutionary arms race between brood parasites and their hosts because they show that the presence of the host's own nestlings for comparison is likely a key clue to favour the evolution of fledgling discrimination and provide new insights on several relevant points such as learning mechanisms and multiparasitism.

Eavesdropping cuckoos: further insights on great spotted cuckoo preference by magpie nests and egg colour.

Reproductive success of brood parasites largely depends on appropriate host selection and, although the use of inadvertent social information emitted by hosts may be of selective advantage for cuckoos, this possibility has rarely been experimentally tested. Here, we manipulated nest size and clutch colouration of magpies (Pica pica), the main host of great spotted cuckoos (Clamator glandarius). These phenotypic traits may potentially reveal information about magpie territory and/or parental quality and could hence influence the cuckoo's choice of host nests. Experimentally reduced magpie nests suffered higher predation rate, and prevalence of cuckoo parasitism was higher in magpie nests with the densest roofs, which suggests a direct advantage for great spotted cuckoos choosing this type of magpie nest. Colouration of magpie clutches was manipulated by adding one artificial egg (blue or cream colouration) at the beginning of the egg-laying period. We found that host nests holding an experimental cream egg experienced a higher prevalence of cuckoo parasitism than those holding a blue-coloured egg. Results from these two experiments suggest that great spotted cuckoos cue on magpie nest characteristics and the appearance of eggs to decide parasitism, and confirm, for the first time, the ability of cuckoos to distinguish between eggs of different colours within the nest of their hosts. Several hypothetical scenarios explaining these results are discussed.

Do climatic conditions affect host and parasite phenotypes differentially? A case study of magpies and great spotted cuckoos.

Climatic conditions, through their effects on resource availability, may affect important life history strategies and trade-offs in animals, as well as their interactions with other organisms such as parasites. This impact may depend on species-specific pathways of development that differ even among species with similar resource requirements (e.g., avian brood parasites and their hosts). Here we explore the degree of covariation between environmental-climatic conditions and nestling phenotypes (i.e., tarsus length, body mass, immune response to phytohemagglutinin injection) and ectoparasite loads of great spotted cuckoos (Clamator glandarius) and those of their magpie (Pica pica) hosts, both within and among 11 study years (1997-2011). Our main results were that (1) nestling phenotypes differed among years, but differently for great spotted cuckoos and magpies; (2) nestling phenotypes showed significant among-year covariation with breeding climatic conditions (temperature and precipitation); and (3) these associations differed for cuckoos and magpies for some phenotypic traits. As the average temperature at the beginning of the breeding season (April) increased, body mass and tarsus length increased only for cuckoos, but not for magpie hosts, while immune response decreased in both species. Finally, (4) the strength of the within-year relationships between the probability of ectoparasitism by Carnus hemapterus flies and laying date (used as an estimate of the within-year variation in climatic conditions) was negatively affected by the annual accumulated precipitation in April. These results strongly suggest that variation in climatic conditions would result in asymmetric effects on different species with respect to the probability of ectoparasitism, immunity and body size. Such asymmetric effects may affect animal interactions in general and those of brood parasites and their hosts in particular.

Brood parasitism correlates with the strength of spatial autocorrelation of life history and defensive traits in Magpies.

Environmental characteristics of neighboring locations are generally more similar than those of distant locations. Selection pressures due to parasitism and other environmental conditions shape life history traits of hosts; thus, the probability of parasitism should be associated with the strength of spatial autocorrelation in life history and defensive traits of their hosts. Here we test this hypothesis in three different subpopulations of Magpie (Pica pica) parasitized by the Great Spotted Cuckoo (Clamator glandarius) during three breeding seasons. In some of the years and study plots, we found evidence of positive spatial autocorrelations for clutch size and parasitism rate, but not for laying date. As predicted, brood parasitism was associated with the strength of these spatial autocorrelations. Magpies that bred close to each other in areas of high risk of parasitism responded similarly to experimental parasitic eggs. Moreover, an elevated risk of parasitism eliminated the spatial autocorrelation for clutch size, which became randomly distributed. We discuss possible mechanisms explaining these associations, which may have important consequences for estimating evolutionary responses of hosts to parasitic infections and, therefore, for epidemiological, ecological, and evolutionary studies of host-parasite relationships.

Intraclutch variation in avian eggshell pigmentation: the anaemia hypothesis.

Many passerine species lay eggs that are speckled with dark protoporphyrin pigmentation. Because protoporphyrin is mainly derived from the blood, we here formulate and test a new hypothesis that links an increase in anaemia along the laying sequence to within-clutch variation in egg pigmentation. More intense pigmentation is expected if pigments accumulate during enhanced red blood cell production in response to anaemia. Reduced pigmentation is expected if pigments are derived from the degradation of red blood cells that circulate in smaller numbers due to blood loss. To test this hypothesis, we manipulated anaemia in great tit (Parus major) females by infesting the nests with hen fleas (Ceratophyllus gallinae) prior to egg laying. Polychromatophil (i.e., immature red blood cells) percentage, as a measure of blood cell production, was positively correlated with parasite load confirming that female great tits experienced stronger anaemia when infested with haematophagous parasites during egg laying. We found a positive relationship between spot darkness and laying order that weakened under high parasite load. This result suggests that anaemia in females due to blood-sucking parasites led to diminished protoporphyrin from disintegrated red blood cells and hence a decreased deposition of protoporphyrin. However, the overall increase in pigment darkness along the laying sequence suggests that pigments also accumulate by enhanced red blood cell production caused by anaemia due to egg production itself.

Corticosterone levels in host and parasite nestlings: is brood parasitism a hormonal stressor?

Parasite chicks from non-evictor species usually try to monopolize host parental care, thereby increasing considerably the level of food competition in the nest. Here, we propose that brood parasitism is an important stressor for host and parasite nestlings and explore this hypothesis in the non-evictor great spotted cuckoo (Clamator glandarius) and its main hosts, the same-sized black-billed magpie (Pica pica) and the larger carrion crow (Corvus corone). We experimentally created 3-nestling broods of different brood compositions (only cuckoo chicks, only host chicks, or cuckoo and host chicks together) and measured baseline corticosterone levels of nestlings along their developmental period (early, middle and late). We found that brood parasitism increased corticosterone levels in magpie nestlings in the mid and late nestling period compared to those raised in unparasitized nests. Interestingly, carrion crow nestlings from parasitized nests only increased their corticosterone levels in the mid nestling period, when the competition for food with the cuckoo nestling was highest. Our results suggest that brood parasitism could be a potential physiological stressor for host nestlings, especially during the developmental stages where food requirements are highest. Conversely, cuckoo nestlings could be physiologically adapted to high competition levels since they did not show significant differences in corticosterone levels in relation to brood composition.

Effects of early developmental conditions on innate immunity are only evident under favourable adult conditions in zebra finches.

Long-term effects of unfavourable conditions during development can be expected to depend on the quality of the environment experienced by the same individuals during adulthood. Yet, in the majority of studies, long-term effects of early developmental conditions have been assessed under favourable adult conditions only. The immune system might be particularly vulnerable to early environmental conditions as its development, maintenance and use are thought to be energetically costly. Here, we studied the interactive effects of favourable and unfavourable conditions during nestling and adult stages on innate immunity (lysis and agglutination scores) of captive male and female zebra finches (Taeniopygia guttata). Nestling environmental conditions were manipulated by a brood size experiment, while a foraging cost treatment was imposed on the same individuals during adulthood. This combined treatment showed that innate immunity of adult zebra finches is affected by their early developmental conditions and varies between both sexes. Lysis scores, but not agglutination scores, were higher in individuals raised in small broods and in males. However, these effects were only present in birds that experienced low foraging costs. This study shows that the quality of the adult environment may shape the long-term consequences of early developmental conditions on innate immunity, as long-term effects of nestling environment were only evident under favourable adult conditions.

Variation in innate immunity in relation to ectoparasite load, age and season: a field experiment in great tits (Parus major).

It remains largely unknown which factors affect the innate immune responses of free-living birds. Nevertheless, the degree of innate immunity may play a crucial role in an individual's survival as it procures the first defence against pathogens. We manipulated the ectoparasite load of great tit (Parus major) nests by infesting them with hen fleas (Ceratophyllus gallinae) before egg laying. We subsequently quantified natural antibody (NAb) concentration and complement activation in nestlings and adult females during breeding and post-breeding periods. NAb concentrations increased in nestlings and adult females breeding in flea-infested nest boxes during the nestling provisioning period, but not in breeding females during incubation. In contrast, parasite abundance did not affect levels of complement activity in females. NAb levels of nestlings were already fully developed at the end of the nestling stage, but complement activation was only observed post-fledging. Concentrations of NAbs and complement activation of adult females were significantly lower during the breeding season compared with post-breeding levels, but did not differ between incubation and chick rearing. Further experimental studies in species that vary in life-history strategies will allow us to unravel the mechanisms underlying the observed variation in innate immune defences.

Kestrel-prey dynamic in a Mediterranean region: the effect of generalist predation and climatic factors.

BACKGROUND: Most hypotheses on population limitation of small mammals and their predators come from studies carried out in northern latitudes, mainly in boreal ecosystems. In such regions, many predators specialize on voles and predator-prey systems are simpler compared to southern ecosystems where predator communities are made up mostly of generalists and predator-prey systems are more complex. Determining food limitation in generalist predators is difficult due to their capacity to switch to alternative prey when the basic prey becomes scarce. METHODOLOGY: We monitored the population density of a generalist raptor, the Eurasian kestrel Falco tinnunculus over 15 years in a mountainous Mediterranean area. In addition, we have recorded over 11 years the inter-annual variation in the abundance of two main prey species of kestrels, the common vole Microtus arvalis and the eyed lizard Lacerta lepida and a third species scarcely represented in kestrel diet, the great white-toothed shrew Crocidura russula. We estimated the per capita growth rate (PCGR) to analyse population dynamics of kestrel and predator species. PRINCIPAL FINDINGS: Multimodel inference determined that the PCGR of kestrels was better explained by a model containing the population density of only one prey species (the common vole) than a model using a combination of the densities of the three prey species. The PCGR of voles was explained by kestrel abundance in combination with annual rainfall and mean annual temperature. In the case of shrews, growth rate was also affected by kestrel abundance and temperature. Finally, we did not find any correlation between kestrel and lizard abundances. SIGNIFICANCE: Our study showed for the first time vertebrate predator-prey relationships at southern latitudes and determined that only one prey species has the capacity to modulate population dynamics of generalist predators and reveals the importance of climatic factors in the dynamics of micromammal species and lizards in the Mediterranean region.

Effects of maternal carotenoid availability in relation to sex, parasite infection and health status of nestling kestrels (Falco tinnunculus).

Mothers can strongly influence the development of their offspring, and if maternal resources are limited, they may influence optimal reproductive strategies. In birds, maternally deposited carotenoids are a prominent component of egg yolk and are vital for the development of the embryo. However, results of long-lasting fitness consequences of this early nutritional environment have been scarce and inconsistent. In addition, sex-biased sensitivity to different egg components is one of the mechanisms postulated to account for sex-linked environmental vulnerability during early life. However, this important aspect is usually not accounted for when investigating maternal investment in carotenoids. In this study we gave carotenoid (lutein) supplements to female Eurasian kestrels (Falco tinnunculus) before and during egg laying. The experiment increased female plasma carotenoids, but this effect was not apparent in hatchling and fledgling plasma carotenoid concentration. Also, results showed that carotenoid supplementation increased the high density lipoprotein to low density lipoprotein ratio in adult females, suggesting that dietary carotenoids may influence lipid metabolism. Furthermore, the effect of the treatment was manifested in several nestling health state parameters. Nestlings of carotenoid-supplemented females were infested by less intestinal parasite groups, had higher lymphocyte concentrations in blood plasma, and were less stressed (heterophile to lymphocyte ratio) than control nestlings. In addition, an interaction between the experimental treatment and nestling sex was apparent for globulin concentrations, favouring the smaller male nestlings. Thereby, suggesting that males benefited more than females from an increase in maternal carotenoid investment. Our study shows that an increase in carotenoids in the maternal diet during egg laying favours nestling development in kestrels, and may also affect nestlings in a sex-specific way.