Immune challenge reduces daily activity period in free-living birds for three weeks.

Non-lethal infections are common in free-living animals and the associated sickness behaviours can impact crucial life-history trade-offs. However, little is known about the duration and extent of such sickness behaviours in free-living animals, and consequently how they affect life-history decisions. Here, free-living Eurasian blackbirds, Turdus merula, were immune-challenged with lipopolysaccharide (LPS) to mimic a bacterial infection and their behaviour was monitored for up to 48 days using accelerometers. As expected, immune-challenged birds were less active than controls within the first 24 h. Unexpectedly, this reduced activity remained detectable for 20 days, before both groups returned to similar activity levels. Furthermore, activity was positively correlated with a pre-experimental index of complement activity, but only in immune-challenged birds, suggesting that sickness behaviours are modulated by constitutive immune function. Differences in daily activity levels stemmed from immune-challenged birds resting earlier at dusk than control birds, while activity levels between groups were similar during core daytime hours. Overall, activity was reduced by 19% in immune-challenged birds and they were on average almost 1 h less active per day for 20 days. This unexpected longevity in sickness behaviour may have severe implications during energy-intense annual-cycle stages (e.g. breeding, migration, winter). Thus, our data help to understand the consequences of non-lethal infections on free-living animals.

Assessment of avian health status: suitability and constraints of the Zoetis VetScan VS2 blood analyser for ecological and evolutionary studies.

Biochemical analyses of blood can decipher physiological conditions of living animals and unravel mechanistic underpinnings of life-history strategies and trade-offs. Yet, researchers in ecology and evolution often face constraints in which methods to apply, not least due to blood volume restrictions or field settings. Here, we test the suitability of a portable biochemical analyser (Zoetis VetScan VS2) for ecological and evolutionary studies that may help solve those problems. Using as little as 80 µl of whole-bird blood from free-living Jackdaws (Corvus monedula) and captive Zebra Finches (Taeniopygia guttata), we show that eight (out of 10) blood analytes show high repeatability after short-term storage (approximately 2 h) and six after 12 h storage time. Handling stress had a clear impact on all except two analytes by 16 min after catching. Finally, six analytes showed consistency within individuals over a period of 30 days, and three even showed individual consistency over a year. Taken together, we conclude that the VetScan VS2 captures biologically relevant variation in blood analytes using just 80 µl of whole blood and, thus, provides valuable physiological measurements of (small) birds sampled in semi-field and field conditions.

Interactive effects of rising temperatures and urbanisation on birds across different climate zones: A mechanistic perspective.

Climate change and urbanisation are among the most pervasive and rapidly growing threats to biodiversity worldwide. However, their impacts are usually considered in isolation, and interactions are rarely examined. Predicting species' responses to the combined effects of climate change and urbanisation, therefore, represents a pressing challenge in global change biology. Birds are important model taxa for exploring the impacts of both climate change and urbanisation, and their behaviour and physiology have been well studied in urban and non-urban systems. This understanding should allow interactive effects of rising temperatures and urbanisation to be inferred, yet considerations of these interactions are almost entirely lacking from empirical research. Here, we synthesise our current understanding of the potential mechanisms that could affect how species respond to the combined effects of rising temperatures and urbanisation, with a focus on avian taxa. We discuss potential interactive effects to motivate future in-depth research on this critically important, yet overlooked, aspect of global change biology. Increased temperatures are a pronounced consequence of both urbanisation (through the urban heat island effect) and climate change. The biological impact of this warming in urban and non-urban systems will likely differ in magnitude and direction when interacting with other factors that typically vary between these habitats, such as resource availability (e.g. water, food and microsites) and pollution levels. Furthermore, the nature of such interactions may differ for cities situated in different climate types, for example, tropical, arid, temperate, continental and polar. Within this article, we highlight the potential for interactive effects of climate and urban drivers on the mechanistic responses of birds, identify knowledge gaps and propose promising future research avenues. A deeper understanding of the behavioural and physiological mechanisms mediating species' responses to urbanisation and rising temperatures will provide novel insights into ecology and evolution under global change and may help better predict future population responses.

Cloacal microbiota are biogeographically structured in larks from desert, tropical and temperate areas.

BACKGROUND: In contrast with macroorganisms, that show well-documented biogeographical patterns in distribution associated with local adaptation of physiology, behavior and life history, strong biogeographical patterns have not been found for microorganisms, raising questions about what determines their biogeography. Thus far, large-scale biogeographical studies have focused on free-living microbes, paying little attention to host-associated microbes, which play essential roles in physiology, behavior and life history of their hosts. Investigating cloacal gut microbiota of closely-related, ecologically similar free-living songbird species (Alaudidae, larks) inhabiting desert, temperate and tropical regions, we explored influences of geographical location and host species on α-diversity, co-occurrence of amplicon sequence variants (ASVs) and genera, differentially abundant and dominant bacterial taxa, and community composition. We also investigated how geographical distance explained differences in gut microbial community composition among larks. RESULTS: Geographic location did not explain variation in richness and Shannon diversity of cloacal microbiota in larks. Out of 3798 ASVs and 799 bacterial genera identified, 17 ASVs (< 0.5%) and 43 genera (5%) were shared by larks from all locations. Desert larks held fewer unique ASVs (25%) than temperate zone (31%) and tropical larks (34%). Five out of 33 detected bacterial phyla dominated lark cloacal gut microbiomes. In tropical larks three bacterial classes were overrepresented. Highlighting the distinctiveness of desert lark microbiota, the relative abundances of 52 ASVs differed among locations, which classified within three dominant and 11 low-abundance phyla. Clear and significant phylogenetic clustering in cloacal microbiota community composition (unweighted UniFrac) showed segregation with geography and host species, where microbiota of desert larks were distinct from those of tropical and temperate regions. Geographic distance was nonlinearly associated with pairwise unweighted UniFrac distances. CONCLUSIONS: We conclude that host-associated microbiota are geographically structured in a group of widespread but closely-related bird species, following large-scale macro-ecological patterns and contrasting with previous findings for free-living microbes. Future work should further explore if and to what extent geographic variation in host-associated microbiota can be explained as result of co-evolution between gut microbes and host adaptive traits, and if and how acquisition from the environmental pool of bacteria contributes to explaining host-associated communities.

Recovery of constitutive immune function after migratory endurance flight in free-living birds.

Strenuous physical activity can negatively affect constitutive innate immune function (CIF), the always present first line of defence against pathogens. CIF is non-specific, and thus vital when encountering novel pathogens. A lowered CIF likely increases the risk of infection and disease. Migratory birds engage in truly extreme physical activity during their endurance flights, however, little is known about how they deal with the negative impact this has on their immune function. By collecting both between- and within-individual data we show, for the first time, that free-flying migratory birds can recover several parameters of CIF during stopovers, which are stationary periods in between migratory flights. With this, we provide an important piece of the puzzle on how migrating birds cope with the physiological challenges they face on their biannual journeys. Furthermore, our study stresses the importance of migratory stopovers beyond fuel accumulation.

Holding time has limited impact on constitutive innate immune function in a long-lived Antarctic seabird, the Adélie penguin: implications for field studies.

There is great interest in measuring immune function in wild animals. Yet, field conditions often have methodological challenges related to handling stress, which can alter physiology. Despite general consensus that immune function is influenced by handling stress, previous studies have provided equivocal results. Furthermore, few studies have focused on long-lived species, which may have different stress-immune trade-offs compared to short-lived species that have primarily been tested. Here, we investigate whether capture and handling duration impacts innate immune function in a long-lived seabird, the Adélie penguin (Pygoscelis adeliae). We found no evidence for changes in three commonly used parameters of innate immune function upon holding time of up to 2 h, suggesting that immune function in this species is more robust against handling than in other species. This opens up exciting possibilities for measuring immune function in species with similar life-histories even if samples cannot be taken directly after capture.

Innate immune function and antioxidant capacity of nestlings of an African raptor covary with the level of urbanisation around breeding territories.

Urban areas provide breeding habitats for many species. However, animals raised in urban environments face challenges such as altered food availability and quality, pollution and pathogen assemblages. These challenges can affect physiological processes such as immune function and antioxidant defences which are important for fitness. Here, we explore how levels of urbanisation influence innate immune function, immune response to a mimicked bacterial infection and antioxidant capacity of nestling Black Sparrowhawks Accipiter melanoleucus in South Africa. We also explore the effect of timing of breeding and rainfall on physiology since both can influence the environmental condition under which nestlings are raised. Finally, because urbanisation can influence immune function indirectly, we use path analyses to explore direct and indirect associations between urbanisation, immune function and oxidative stress. We obtained measures of innate immunity (haptoglobin, lysis, agglutination, bactericidal capacity), indices of antioxidant capacity (total non-enzymatic antioxidant capacity (tAOX) and total glutathione from nestlings from 2015 to 2019. In addition, in 2018 and 2019, we mimicked a bacterial infection by injecting nestlings with lipopolysaccharide and quantified their immune response. Increased urban cover was associated with an increase in lysis and a decrease in tAOX, but not with any of the other physiological parameters. Furthermore, except for agglutination, no physiological parameters were associated with the timing of breeding. Lysis and bactericidal capacity, however, varied consistently with the annual rainfall pattern. Immune response to a mimicked a bacterial infection decreased with urban cover but not with the timing of breeding nor rainfall. Our path analyses suggested indirect associations between urban cover and some immune indices via tAOX but not via the timing of breeding. Our results show that early-life development in an urban environment is associated with variation in immune and antioxidant functions. The direct association between urbanisation and antioxidant capacity and their impact on immune function is likely an important factor mediating the impact of urbanisation on urban-dwelling animals. Future studies should explore how these results are linked to fitness and whether the responses are adaptive for urban-dwelling species.

Telomeres in ecology and evolution: A review and classification of hypotheses.

Research on telomeres in the fields of ecology and evolution has been rapidly expanding over the last two decades. This has resulted in the formulation of a multitude of, often name-given, hypotheses related to the associations between telomeres and life-history traits or fitness-facilitating processes (and the mechanisms underlying them). However, the differences (or similarities) between the various hypotheses, which can originate from different research fields, are often not obvious. Our aim here is therefore to give an overview of the hypotheses that are of interest in ecology and evolution and to provide two frameworks that help discriminate among them. We group the hypotheses (i) based on their association with different research questions, and (ii) using a hierarchical approach that builds on the assumptions they make, such as about causality of telomere length/shortening and/or the proposed functional consequences of telomere shortening on organism performance. Both our frameworks show that there exist parallel lines of thoughts in different research fields. Moreover, they also clearly illustrate that there are in many cases competing hypotheses within clusters, and that some of these even have contradictory assumptions and/or predictions. We also touch upon two topics in telomere research that would benefit from further conceptualization. This review should help researchers, both those familiar with and those new to the subject, to identify future avenues of research.

Corrigendum to: Not just fuel: energy stores are correlated with immune function and oxidative damage in a long-distance migrant.

[This corrects the article DOI: 10.1093/cz/zoz009.][This corrects the article DOI: 10.1093/cz/zoz009.].

Reduced ectoparasite load, body mass and blood haemolysis in Eurasian kestrels (Falco tinnunculus) along an urban-rural gradient.

Urbanisation is proceeding at an alarming rate which forces wildlife to either retreat from urban areas or cope with novel stressors linked to human presence and activities. For example, urban stressors like anthropogenic noise, artificial light at night and chemical pollution can have severe impacts on the physiology of wildlife (and humans), in particular the immune system and antioxidant defences. These physiological systems are important to combat and reduce the severity of parasitic infections, which are common among wild animals. One question that then arises is whether urban-dwelling animals, whose immune and antioxidant system are already challenged by the urban stressors, are more susceptible to parasitic infections. To assess this, we studied nestlings of Eurasian kestrels (Falco tinnunculus) in Vienna, Austria, during 2015 and 2017. We measured biomarkers of innate immune function, oxidative stress and body mass index and ectoparasite infection intensity in 143 nestlings (from 56 nests) along an urban gradient. Nestlings in more urbanised areas had overall fewer ectoparasites, lower haemolysis (complement activity) and lower body mass index compared to nestlings in less urbanised areas. None of the other immune or oxidative stress markers were associated with the urban gradient. Despite some non-significant results, our data still suggest that kestrel nestlings experience some level of reduced physiological health, perhaps as a consequence of exposure to more urban stressors or altered prey availability in inner-city districts even though they had an overall lower ectoparasite burden in these heavily urbanised areas.

Parental morph combination does not influence innate immune function in nestlings of a colour-polymorphic African raptor.

Conditions experienced during early life can have long-term individual consequences by influencing dispersal, survival, recruitment and productivity. Resource allocation during development can have strong carry-over effects onto these key parameters and is directly determined by the quality of parental care. In the black sparrowhawk (Accipiter melanoleucus), a colour-polymorphic raptor, parental morphs influence nestling somatic growth and survival, with pairs consisting of different colour morphs ('mixed-morph pairs') producing offspring with lower body mass indices, but higher local apparent survival rates. Resource allocation theory could explain this relationship, with nestlings of mixed-morph pairs trading off a more effective innate immune system against somatic growth. We quantified several innate immune parameters of nestlings (hemagglutination, hemolysis, bacteria-killing capacity and haptoglobin concentration) and triggered an immune response by injecting lipopolysaccharides. Although we found that nestlings with lower body mass index had higher local survival rates, we found no support for the proposed hypothesis: neither baseline immune function nor the induced immune response of nestlings was associated with parental morph combination. Our results suggest that these immune parameters are unlikely to be involved in providing a selective advantage for the different colour morphs' offspring, and thus innate immunity does not appear to be traded off against a greater allocation of resources to somatic growth. Alternative hypotheses explaining the mechanism of a low nestling body mass index leading to subsequent higher local survival could be related to the post-fledgling dependency period or differences in dispersal patterns for the offspring from different morph combinations.

Exposure to artificial light at night alters innate immune response in wild great tit nestlings.

The large-scale impact of urbanization on wildlife is rather well documented; however, the mechanisms underlying the effects of urban environments on animal physiology and behaviour are still poorly understood. Here, we focused on one major urban pollutant - artificial light at night (ALAN) - and its effects on the capacity to mount an innate immune response in wild great tit (Parus major) nestlings. Exposure to ALAN alters circadian rhythms of physiological processes, by disrupting the nocturnal production of the hormone melatonin. Nestlings were exposed to a light source emitting 3 lx for seven consecutive nights. Subsequently, nestlings were immune challenged with a lipopolysaccharide injection, and we measured haptoglobin and nitric oxide levels pre- and post-injection. Both haptoglobin and nitric oxide are important markers for innate immune function. We found that ALAN exposure altered the innate immune response, with nestlings exposed to ALAN having lower haptoglobin and higher nitric oxide levels after the immune challenge compared with dark-night nestlings. Unexpectedly, nitric oxide levels were overall lower after the immune challenge than before. These effects were probably mediated by melatonin, as ALAN-treated birds had on average 49% lower melatonin levels than the dark-night birds. ALAN exposure did not have any clear effects on nestling growth. This study provides a potential physiological mechanism underlying the documented differences in immune function between urban and rural birds observed in other studies. Moreover, it gives evidence that ALAN exposure affects nestling physiology, potentially causing long-term effects on physiology and behaviour, which ultimately can affect their fitness.

Jackdaw nestlings rapidly increase innate immune function during the nestling phase but no evidence for a trade-off with growth.

Although animals are born with a protective immune system, even the innate immune system is under development from birth to adulthood and this development may be affected by sex and growth. However, most knowledge comes from captive animals or long-lived slow growing species. Moreover, little is known about how innate immune function, the important first line of defence, develops during early life in fast-growing animals such as free-living passerines. We studied development of innate baseline immune function in nestlings of free-living jackdaws Corvus monedula. We measured four immune parameters (hemolysis, hemagglutination, bacterial-killing capacity, haptoglobin concentration) and structural body size (body mass, wing length, tarsus length) at day 12 and day 29 post-hatching. We found that three out of four immune parameters (hemolysis, hemagglutination, bacterial-killing capacity) substantially increased with nestling age and had roughly reached adult levels shortly prior to fledging. We found little differences in immune development between males and females despite them differing in structural development. We also found no evidence that the nestlings traded off immune development with growth. That nestlings rapidly increase innate baseline immune function during early life and similarly in males and females indicates the importance of a well-functioning immune system already during the nestling phase.

A Comparative Study of the Innate Humoral Immune Response to Avian Influenza Virus in Wild and Domestic Mallards.

Domestic mallards (Anas platyrhynchos domesticus) are traditionally used as a model to investigate infection dynamics and immune responses to low pathogenic avian influenza viruses (LPAIVs) in free-living mallards. However, it is unclear whether the immune response of domestic birds reflects the response of their free-living counterparts naturally exposed to these viruses. We investigated the extent to which the innate humoral immune response was similar among (i) wild-type domestic mallards in primary and secondary infection with LPAIV H4N6 in a laboratory setting (laboratory mallards), (ii) wild-type domestic mallards naturally exposed to LPAIVs in a semi-natural setting (sentinel mallards), and (iii) free-living mallards naturally exposed to LPAIVs. We quantified innate humoral immune function by measuring non-specific natural antibodies (agglutination), complement activity (lysis), and the acute phase protein haptoglobin. We demonstrate that complement activity in the first 3 days after LPAIV exposure was higher in primary-exposed laboratory mallards than in sentinel and free-living mallards. LPAIV H4N6 likely activated the complement system and the acute phase response in primary-exposed laboratory mallards, as lysis was higher and haptoglobin lower at day 3 and 7 post-exposure compared to baseline immune function measured prior to exposure. There were no differences observed in natural antibody and haptoglobin concentrations among laboratory, sentinel, and free-living mallards in the first 3 days after LPAIV exposure. Our study demonstrates that, based on the three innate humoral immune parameters measured, domestic mallards seem an appropriate model to investigate innate immunology of their free-living counterparts, albeit the innate immune response of secondary-LPAIV exposed mallards is a better proxy for the innate immune response in pre-exposed free-living mallards than that of immunologically naïve mallards.

Not just fuel: energy stores are correlated with immune function and oxidative damage in a long-distance migrant.

In many animals, catabolic and anabolic periods are temporally separated. Migratory birds alternate energy expenditure during flight with energy accumulation during stopover. The size of the energy stores at stopover affects the decision to resume migration and thus the temporal organization of migration. We now provide data suggesting that it is not only the size of the energy stores per se that may influence migration scheduling, but also the physiological consequences of flying. In two subspecies of the northern wheatear Oenanthe oenanthe, a long-distance migrant, estimated energy stores at a stopover during autumn migration were positively related with both constitutive innate and acquired immune function, and negatively related with oxidative damage to lipids. In other words, migrants' physiological condition was associated with their energetic condition. Although time spent at stopover before sampling may have contributed to this relationship, our results suggest that migrants have to trade-off the depletion of energy stores during flight with incurring physiological costs. This will affect migrants' decisions when to start and when to terminate a migratory flight. The physiological costs associated with the depletion of energy stores may also help explaining why migrants often arrive at and depart from stopover sites with larger energy stores than expected. We propose that studies on the role of energy stores as drivers of the temporal organization of (avian) migration need to consider physiological condition, such as immunological and oxidative states.

Reduced immune responsiveness contributes to winter energy conservation in an Arctic bird.

Animals in seasonal environments must prudently manage energy expenditure to survive the winter. This may be achieved through reductions in the allocation of energy for various purposes (e.g. thermoregulation, locomotion, etc.). We studied whether such trade-offs also include suppression of the innate immune response, by subjecting captive male Svalbard ptarmigan (Lagopus muta hyperborea) to bacterial lipopolysaccharide (LPS) during exposure to either mild temperature (0°C) or cold snaps (acute exposure to -20°C), in constant winter darkness when birds were in energy-conserving mode, and in constant daylight in spring. The innate immune response was mostly unaffected by temperature. However, energy expenditure was below baseline when birds were immune challenged in winter, but significantly above baseline in spring. This suggests that the energetic component of the innate immune response was reduced in winter, possibly contributing to energy conservation. Immunological parameters decreased (agglutination, lysis, bacteriostatic capacity) or did not change (haptoglobin/PIT54) after the challenge, and behavioural modifications (anorexia, mass loss) were lengthy (9 days). While we did not study the mechanisms explaining these weak, or slow, responses, it is tempting to speculate they may reflect the consequences of having evolved in an environment where pathogen transmission rate is presumably low for most of the year. This is an important consideration if climate change and increased exploitation of the Arctic would alter pathogen communities at a pace outwith counter-adaption in wildlife.

Migrating birds rapidly increase constitutive immune function during stopover.

Migratory flight is physiologically highly demanding and has been shown to negatively affect multiple parameters of constitutive immune function (CIF), an animal's first line of physiological defence against infections. In between migratory flights, most birds make stopovers, periods during which they accumulate fuel for the next flight(s). Stopovers are also commonly thought of as periods of rest and recovery, but what this encompasses is largely undefined. Here, we show that during stopover, northern wheatears Oenanthe oenanthe, a long-distance migratory bird, can rapidly increase constitutive innate immune function. We caught and temporarily caged birds under ad libitum food conditions at a stopover site in autumn. Within 2 days, most birds significantly increased complement activity and their ability to kill microbes. Changes in immune function were not related to the birds' food intake or extent of fuel accumulation. Our study suggests that stopovers may not only be important to refuel but also to restore immune function. Additionally, the increase in CIF could help migrating birds to deal with novel pathogens they may encounter at stopover sites.

Experimental facilitation of heat loss affects work rate and innate immune function in a breeding passerine bird.

The capacity to get rid of excess heat produced during hard work is a possible constraint on parental effort during reproduction [heat dissipation limit (HDL) theory]. We released hard-working blue tits (Cyanistes caeruleus) from this constraint by experimentally removing ventral plumage. We then assessed whether this changed their reproductive effort (feeding rate and nestling size) and levels of self-maintenance (change in body mass and innate immune function). Feather-clipped females reduced the number of feeding visits and increased levels of constitutive innate immunity compared with unclipped females but did not fledge smaller nestlings. Thus, they increased self-maintenance without compromising current reproductive output. In contrast, feather clipping did not affect the number of feeding visits or innate immune function in males, despite increased heat loss rate. Our results show that analyses of physiological parameters, such as constitutive innate immune function, can be important when trying to understand sources of variation in investment in self-maintenance versus reproductive effort and that risk of overheating can influence innate immune function during reproduction.

Defence versus defence: Are crucian carp trading off immune function against predator-induced morphology?

Numerous species adopt inducible defence strategies; that is, they have phenotypically plastic traits that decrease the risk of capture and consumption by potential predators. The benefits of expressing alternative phenotypes in high- vs. low-risk environments are well documented. However, inducible anti-predator traits are also expected to incur costs, as they are not expressed when predators are absent, yet empirical evidence of such costs remains scarce. Virtually, all animals in nature are simultaneously under strong selection to evade both capture by predators and infection by parasites or pathogens and, hence, display a diverse arsenal of defences to combat these threats, raising the possibility of trade-offs between defences. A classic example of a predator-induced morphological defence is the deep-bodied shape of crucian carp that reduces risk of predation from gape-size-limited predators. The goal of this study was to examine whether predator exposure affects also immune function in crucian carp, and whether the degree of expressed morphological defence is traded off against immune function in individuals. Following exposure to manipulations of perceived risk (predator presence/absence) in a long-term experiment (8 months), key aspects of innate immune function and individual differences in the expression of inducible morphological defence were quantified. Predator-exposed individuals showed lower haptoglobin levels and complement activity, but higher natural antibody titres than fish from predator-free conditions. When experimentally challenged with a mimicked bacterial infection (LPS injection), fish reared in the presence of a natural predator showed a weaker immune response. Moreover, among predator-exposed individuals, the magnitude of morphological defence expression correlated with both baseline immune function and the ability to mount an immune response. However, these relationships were not consistently supportive of a general trade-off among defences. Our results suggest that fish exposed to predators on average reduce investment in immune function, and, further, the observed relationships among defences in predator-exposed individuals can best be explained from individual fitness and pace-of-life perspectives.

Immune function and blood parasite infections impact stopover ecology in passerine birds.

Stopovers play a crucial role for the success of migrating animals and are key to optimal migration theory. Variation in refuelling rates, stopover duration and departure decisions among individuals has been related to several external factors. The physiological mechanisms shaping stopover ecology are, however, less well understood. Here, we explore how immune function and blood parasite infections relate to several aspects of stopover behaviour in autumn migrating short- and long-distance migrating songbirds. We blood sampled individuals of six species and used an automated radio-telemetry system in the stopover area to subsequently quantify stopover duration, 'bush-level' activity patterns (~ 0.1-30 m), landscape movements (~ 30-6000 m), departure direction and departure time. We show that complement activity, the acute phase protein haptoglobin and blood parasite infections were related to prolonged stopover duration. Complement activity (i.e., lysis) and total immunoglobulins were negatively correlated with bush-level activity patterns. The differences partly depended on whether birds were long-distance or short-distance migrants. Birds infected with avian malaria-like parasites showed longer landscape movements during the stopover than uninfected individuals, and birds with double blood parasite infections departed more than 2.5 h later after sunset/sunrise suggesting shorter flight bouts. We conclude that variation in baseline immune function and blood parasite infection status affects stopover ecology and helps explain individual variation in stopover behaviour. These differences affect overall migration speed, and thus can have significant impact on migration success and induce carry-over effects on other annual-cycle stages. Immune function and blood parasites should, therefore, be considered as important factors when applying optimal migration theory.