Daniel R. Ardia

Carotenoids, Immunocompetence, and the Information Content of Sexual Colors: An Experimental Test

Many male birds use carotenoid pigments to acquire brilliant colors that advertise their health and condition to prospective mates. The direct means by which the most colorful males achieve superior health has been debated, however. One hypothesis, based on studies of carotenoids as antioxidants in humans and other animals, is that carotenoids directly boost the immune system of colorful birds. We studied the relationship between carotenoid pigments, immune function, and sexual coloration in zebra finches (Taeniopygia guttata), a species in which males incorporate carotenoid pigments into their beak to attract mates. We tested the hypotheses that increased dietary carotenoid intake enhances immunocompetence in male zebra finches and that levels of carotenoids circulating in blood, which also determine beak coloration, directly predict the immune response of individuals. We experimentally supplemented captive finches with two common dietary carotenoid pigments (lutein and zeaxanthin) and measured cell‐mediated and humoral immunity a month later. Supplemented males showed elevated blood‐carotenoid levels, brighter beak coloration, and increased cell‐mediated and humoral immune responses than did controls. Cell‐mediated responses were predicted directly by changes in beak color and plasma carotenoid concentration of individual birds. These experimental findings suggest that carotenoid‐based color signals in birds may directly signal male health via the immunostimulatory action of ingested and circulated carotenoid pigments.

The design of artificial nestboxes for the study of secondary hole-nesting birds: a review of methodological inconsistencies and potential biases

Abstract. The widespread use of artificial nestboxes has led to significant advances in our knowledge of the ecology, behaviour and physiology of cavity nesting birds, especially small passerines. Nestboxes have made it easier to perform routine monitoring and experimental manipulation of eggs or nestlings, and also repeatedly to capture, identify and manipulate the parents. However, when comparing results across study sites the use of nestboxes may also introduce a potentially significant confounding variable in the form of differences in nestbox design amongst studies, such as their physical dimensions, placement height, and the way in which they are constructed and maintained. However, the use of nestboxes may also introduce an unconsidered and potentially significant confounding variable due to differences in nestbox design amongst studies, such as their physical dimensions, placement height, and the way in which they are constructed and maintained. Here we review to what extent the characteristics of artificial nestboxes (e.g. size, shape, construction material, colour) are documented in the ‘methods’ sections of publications involving hole-nesting passerine birds using natural or excavated cavities or artificial nestboxes for reproduction and roosting. Despite explicit previous recommendations that authors describe in detail the characteristics of the nestboxes used, we found that the description of nestbox characteristics in most recent publications remains poor and insufficient. We therefore list the types of descriptive data that should be included in the methods sections of relevant manuscripts and justify this by discussing how variation in nestbox characteristics can affect or confound conclusions from nestbox studies. We also propose several recommendations to improve the reliability and usefulness of research based on long-term studies of any secondary hole-nesting species using artificial nestboxes for breeding or roosting.

Experimental cooling during incubation leads to reduced innate immunity and body condition in nestling tree swallows

Nest microclimate can have strong effects that can carry over to later life-history stages. We experimentally cooled the nests of tree swallows (Tachycineta bicolor). Females incubating in cooled nests reduced incubation time and allowed egg temperatures to drop, leading to extended incubation periods. We partially cross-fostered nestlings to test carry-over effects of cooling during incubation on nestling innate constitutive immunity, assessed through bacteria killing ability (BKA) of blood. Nestlings that had been cooled as eggs showed a lower ability to kill bacteria than control nestlings, regardless of the treatment of their foster mother. However, there was no effect of treatment of rearing females on nestling BKA in control nestlings, even though cooled females made significantly fewer feeding visits than did control females. This suggests that the effect of cooling occurred during incubation and was not due to carry-over effects on nestling condition. Nestlings that were exposed to experimental cooling as embryos had lower residual body mass and absolute body mass at all four ages measured. Our results indicate that environmental conditions and trade-offs experienced during one stage of development can have important carry-over effects on later life-history stages.

TREE SWALLOWS TRADE OFF IMMUNE FUNCTION AND REPRODUCTIVE EFFORT DIFFERENTLY ACROSS THEIR RANGE

Latitudinal differences in life histories are believed to be underlain by dif- ferences in trade-offs between current and future reproduction. I report differences in trade- offs between parent and offspring across the range of a widespread avian species, the Tree Swallow (Tachycineta bicolor). I manipulated parental effort and found that in Alaska, where yearly adult return rates are low, breeding females increase their reproductive in- vestment to maintain offspring quality, whereas in Tennessee, where yearly adult return rates are high, breeding females are either unable or unwilling to increase reproductive investment and, consequently, raise offspring of lower quality. I further investigated a critical mechanism of self-maintenance that may underlie differences in survival among sites: immunocompetence. Females breeding in Alaska mounted weaker immune responses when raising enlarged broods, whereas females in Tennessee did not, further suggesting a strategy in Alaskan females to incur costs (i.e., reduce future reproduction) in order to maintain offspring quality. Insect availability increased as the breeding season progressed in both sites, but more food was available in Alaska than in Tennessee. This is among the first studies to report geographic differences in immune function consistent with life history theory, which predicts that individuals with higher survival probabilities should invest more in self-maintenance.

Reproductive effort reduces long-term immune function in breeding tree swallows (Tachycineta bicolor)

We examined whether strategies of reproductive allocation may reduce long-term immunocompetence through the effects of manipulated effort on secondary or acquired immunity. We tested whether increased reproductive effort leads to reduced immune function and survival by manipulating brood size in tree swallows (Tachycineta bicolor) and exposing breeding females to a primary and secondary exposure of sheep red blood cells to elicit a humoral immune response. Females raising enlarged broods produced fewer secondary antibodies than did females raising control or reduced broods. Most importantly, individuals with high secondary responses were more likely to survive to breed 3 years after brood manipulations, suggesting that differences in disease susceptibility may be caused by trade–offs in reproductive allocation. We also found that individual quality, measured by clutch initiation date, mediated the effects of brood manipulations, with higher–quality birds showing a greater ability to deal with increases in effort.

Experimental heating reveals nest temperature affects nestling condition in tree swallows (Tachycineta bicolor)

Investment in one life-history stage can have delayed effects on subsequent life-history stages within a single reproductive bout. We experimentally heated tree swallow (Tachycineta bicolor) nests during incubation to test for effects on parental and nestling conditions. Females incubating in heated boxes maintained higher body condition and fed nestlings at higher rates. We cross-fostered nestlings and found that young nestlings (4–7 days old) incubated in heated nests had higher body condition and body mass, regardless of treatment status of their rearing parent. However, older nestlings which were fed by heated females maintained higher condition and body mass regardless of treatment status of their incubating parent. These results indicate that investment in one life-history stage can have multiple pathways of carry-over effects on future life-history stages.

Do carotenoids buffer testosterone-induced immunosuppression? An experimental test in a colourful songbird

Testosterone (T) is hypothesized to be an important honesty reinforcer of animal sexual signals. Owing to its immunosuppressive effects, only those individuals that can immunologically withstand high T levels can develop the most exaggerated traits. To date, few studies have isolated phenotypic or genotypic buffers that provide ‘high-quality’ animals with such an advantage. Dietary carotenoid pigments may in fact confer such a benefit because when in high supply carotenoids boost immunocompetence and coloration in animals like birds and fishes. We examined the experimental effect of T elevation on carotenoid and immune status in male and female zebra finches (Taeniopygia guttata) and found that T was immunostimulatory in a generalized cell-mediated challenge. We also detected a significant interaction between T treatment and the change in plasma carotenoids that occurred during the immune challenge; the relationship between blood carotenoid change and immunity was positive in controls and negative in T-implanted birds. This suggests that, while correlationally birds with high carotenoid stores were inherently better at mounting strong immune responses, experimentally administered T induced birds to deplete carotenoids for maximizing their health. Our findings highlight a nutrient-specific mechanism by which animals escape high immune costs of T elevation and thus can still elevate ornamentation.

Short-term exposure to testosterone propionate leads to rapid bill color and dominance changes in zebra finches

Testosterone (T) can influence both male-male competition and mate choice displays. In zebra finches, female mate choice is based in part on bill color, and bill color has been shown to be enhanced by long-term testosterone supplementation. However, it is not clear whether bill color plays a role in male-male interactions and how bill color responds to shorter-term changes in T. We tested whether a single injection of testosterone propionate (TP) would influence male-male dominance interactions and lead to rapid (over a three-day period) changes in bill color. In addition, we tested whether bill color predicted aggression and dominance. We allowed birds in triads to establish hierarchies and then injected either dominant or subordinate individuals with TP, in addition to establishing sham control triads. We found that red chroma, but not hue, predicted aggressiveness of males. Exposure to TP led both dominant and subordinate birds to increase dominance scores over three days, longer than the <24h period in which injected TP stays active. In addition, exposure to TP increased red chroma and hue in three days showing the dynamic nature of allocation of pigments to the bill. Our results suggest that zebra finches can modulate T and bill color levels over short time periods and these changes may occur through positive feedback between T-levels and dominance.

Thermal Sensitivity of Immune Function: Evidence against a Generalist-Specialist Trade-Off among Endothermic and Ectothermic Vertebrates

Animal body temperature (Tbody) varies over daily and annual cycles, affecting multiple aspects of biological performance in both endothermic and ectothermic animals. Yet a comprehensive comparison of thermal performance among animals varying in Tbody (mean and variance) and heat production is lacking. Thus, we examined the thermal sensitivity of immune function (a crucial fitness determinant) in Vertebrata, a group encompassing species of varying thermal biology. Specifically, we investigated temperature-related variation in two innate immune performance metrics, hemagglutination and hemolysis, for 13 species across all seven major vertebrate clades. Agglutination and lysis were temperature dependent and were more strongly related to the thermal biology of species (e.g., mean Tbody) than to the phylogenetic relatedness of species, although these relationships were complex and frequently surprising (e.g., heterotherms did not exhibit broader thermal performance curves than homeotherms). Agglutination and lysis performance were positively correlated within species, except in taxa that produce squalamine, a steroidal antibiotic that does not lyse red blood cells. Interestingly, we found the antithesis of a generalist-specialist trade-off: species with broader temperature ranges of immune performance also had higher peak performance levels. In sum, we have uncovered thermal sensitivity of immune performance in both endotherms and ectotherms, highlighting the role that temperature and life history play in immune function across Vertebrata.

Immunoregulatory activity of different dietary carotenoids in male zebra finches

Many animals use carotenoid pigments to color their integument and become sexually attractive. These colorants can also serve physiological functions, protecting cells and tissues from oxidative damage as well as stimulating the immune system. Because animals often acquire several different carotenoid pigments from their diet, there is the potential for different carotenoids to exhibit different free-radical-scavenging or immunoenhancing activity. We experimentally tested how two common dietary xanthophylls - lutein and zeaxanthin - may differentially affect the immune system in male zebra finches (Taeniopygia guttata). Male T. guttata derive their red sexual beak colorants from these two carotenoids, and prior studies with this species have shown that lutein and zeaxanthin together boost cell-mediated immunity. We experimentally elevated these two dietary carotenoids separately in two groups of non-breeding zebra finches, but found that lutein-supplemented and zeaxanthin-supplemented males mounted similar cell-mediated immune responses (to phytohaemagglutinin, or PHA). Although zeaxanthin is a more conjugated carotenoid than lutein and has the potential to be a more potent antioxidant, our study suggests that such a subtle structural difference between these two biochemicals does not differentially affect immune performance in this songbird.Summary.Many animals use carotenoid pigments to color their integument and become sexually attractive. These colorants can also serve physiological functions, protecting cells and tissues from oxidative damage as well as stimulating the immune system. Because animals often acquire several different carotenoid pigments from their diet, there is the potential for different carotenoids to exhibit different free-radical-scavenging or immunoenhancing activity. We experimentally tested how two common dietary xanthophylls - lutein and zeaxanthin - may differentially affect the immune system in male zebra finches (Taeniopygia guttata). Male T. guttata derive their red sexual beak colorants from these two carotenoids, and prior studies with this species have shown that lutein and zeaxanthin together boost cell-mediated immunity. We experimentally elevated these two dietary carotenoids separately in two groups of non-breeding zebra finches, but found that lutein-supplemented and zeaxanthin-supplemented males mounted similar cell-mediated immune responses (to phytohaemagglutinin, or PHA). Although zeaxanthin is a more conjugated carotenoid than lutein and has the potential to be a more potent antioxidant, our study suggests that such a subtle structural difference between these two biochemicals does not differentially affect immune performance in this songbird.