Patterns of stress response to foreign eggs by a rejecter host of an obligate avian brood parasite.

One of the most effective defenses of avian hosts against obligate brood parasites is the ejection of parasitic eggs from the nests. Despite the clear fitness benefits of this behavior, individuals within so-called "egg-rejecter" host species still show substantial variation in their propensity to eliminate foreign eggs from the nest. We argue that this variation can be further understood by studying the physiological mechanisms of host responses to brood parasitic egg stimuli: independent lines of research increasingly support the hypothesis that stress-related physiological response to parasitic eggs may trigger egg rejection. The "stress-mediated egg rejection" hypothesis requires that hosts activate the stress-response when responding to parasitic egg stimuli. We tested this prediction by asking whether hosts showed differential stress response when exposed to host-like (mimetic) or parasite-like (non-mimetic) eggs. We experimentally parasitized incubating American robins Turdus migratorius, a robust egg-rejecter host to obligate brood parasitic brown-headed cowbirds Molothrus ater, with mimetic or non-mimetic model eggs. To assess the stress response, we measured the heart rate in incubating females immediately after experimental parasitism. We also measured plasma corticosterone and, in a subset of birds, used RNA-sequencing to analyze the expression of proopiomelanocortin (POMC), a precursor of adrenocorticotropic hormone, 2 h after experimental parasitism. We found that egg type had no effect on heart rate. Two hours following experimental parasitism, plasma corticosterone did not differ between the differently-colored model egg treatments or between rejecter and accepter females within the non-mimetic treatment. However, females exposed to non-mimetic eggs showed an upregulation of POMC gene expression (before FDR correction) in the pituitary compared with females treated with mimetic eggs. Our findings suggest that in an egg-rejecter host species, non-mimetic parasitic eggs may increase the activity of the stress-related hypothalamic-pituitary-adrenal axis compared with mimetic eggs, although the temporal dynamics of this response are not yet understood.

Non-invasive elevation of circulating corticosterone increases the rejection of foreign eggs in female American robins (Turdus migratorius).

Avian obligate brood parasites rely on other species to raise their offspring. In turn, many brood parasite hosts have evolved defensive behaviors to reduce the costs of brood parasitism, yet the proximate bases underlying these defenses remain poorly understood. Recent studies regarding the potential endocrine mechanisms of foreign-egg rejection have implicated corticosterone as a physiological mediator of anti-parasitic defenses. For example, corticosterone is elevated in response to non-mimetic eggs in an egg rejecter thrush, the Eurasian blackbird (Turdus merula) and this hormone's suppression reduces egg rejection rates in the congeneric American robin (T. migratorius). American robins are also among the few host species of obligate brood parasitic brown-headed cowbirds (Molothrus ater) that readily reject foreign eggs from their nests. We non-invasively elevated corticosterone levels in incubating female robins by dissolving it in DMSO gel which was then applied onto eggs already in the clutch. Relative to controls treated with pure DMSO gel, corticosterone-treated female robins were more likely to reject a non-mimetic, cowbird-sized foreign egg (72 %) than control females (50 %) when accounting for the known effect of lower clutch sizes on greater egg rejection. Future studies are needed to assess the sensory and cognitive impact(s) of corticosterone, as well as other hormones essential for parental care, in this and other hosts' defense behaviors against avian brood parasitism.

Ground nesting by arboreal American robins (Turdus migratorius).

Animals with dependent and vulnerable young need to decide where to raise their offspring to minimize ill effects of weather, competition, parasitism, and predation. These decisions have critical fitness consequences through impacting the survival of both adults and progeny. Birds routinely place their nest in specific sites, allowing species to be broadly classified based on nest location (e.g., ground- or tree-nesting). However, from 2018 to 2020, we observed 24 American robin (Turdus migratorius) nests placed not on their species-typical arboreal substrates or human-made structures but on the ground at a predator-rich commercial tree-farm in Illinois, USA. This behavior does not appear to be in response to competition and did not affect nest daily survival rate but was restricted to the early half of the breeding season. We hypothesize that ground nesting may be an adaptive response to avoid exposure and colder temperatures at sites above the ground early in the breeding season or a nonadaptive consequence of latent robin nest-placement flexibility.

Endocrine regulation of egg rejection in an avian brood parasite host.

Parasite-host coevolution can lead to novel behavioural adaptations in hosts to resist parasitism. In avian obligate brood parasite and host systems, many host species have evolved diverse cognitive and behavioural traits to recognize and reject parasitic eggs. Our understanding of the evolution and ecology of these defences hinges on identifying the mechanisms that regulate them. We hypothesized that corticosterone, a hormone linked to stress response, vigilance and the suppression of parental behaviour, stimulates the rejection of foreign eggs by brood parasite hosts. We experimentally reduced circulating glucocorticoid levels with mitotane injections in American robins Turdus migratorius and found that the mitotane-treated birds rejected foreign eggs at a lower frequency compared to the sham-treated subjects. This is the first study to causally identify a potential mechanism of a widespread defence behaviour, and it is consistent with egg rejection being mediated by stress physiology.

Proximate predictors of variation in egg rejection behavior by hosts of avian brood parasites.

The rejection of parasitic eggs by hosts of avian brood parasites is one of the most common and effective defenses against parasitism. Despite its adaptive significance, egg rejection often shows substantial intraspecific variation: some individuals are more likely to remove or abandon parasitic eggs than others. Understanding variation in egg rejection requires that we study factors linked to the ability to perceive parasitic eggs as well as factors that may influence the rejection of a foreign egg once it has been recognized. Here, we asked what cognitive, physiological, and life-history factors explained variation in the rejection of model eggs by American robin (Turdus migratorius) females. We found that the probability of egg rejection was related to the clutch size at the time of parasitism: In support of Weber's law, females with fewer eggs were more likely to reject the model eggs. In turn, females with greater mass and higher corticosterone levels were less likely to reject eggs, and egg rejection probability was negatively related to incubation progress. Our data thus suggest that proximate predictors of an individual's egg rejection behavior include the cognitive environment of the nest, life-history factors, as well as the physiological state of the subject. However, much of the statistical variation in the egg rejection responses of robins to the model eggs remained unexplained. Future experiments should aim to test the causal roles of these and other factors in generating within- and among-individual variation in the rejection of parasitic eggs. (PsycInfo Database Record (c) 2020 APA, all rights reserved).

Fitting different visual models to behavioral patterns of parasitic egg rejection along a natural egg color gradient in a cavity-nesting host species.

Avian brood parasites lay their eggs in other birds' nests, and hosts can mitigate the fitness cost of raising unrelated offspring by rejecting parasitic eggs. A visually-based cognitive mechanism often thought to be used by hosts to discriminate the foreign egg is to compare it against the hosts' own eggshell by size, shape, maculation, and/or ground coloration (i.e., absolute chromatic contrast). However, hosts may instead discriminate eggs based on their colors along a scale of natural avian eggshell coloration (i.e., directional chromatic contrast). In support of this latter visual process, recent research has found that directional chromatic contrasts can explain some host species' rejection behavior better than absolute chromatic or achromatic contrasts. Here, for the first time, we conducted an experiment in a cavity-nesting host species to test the predictions of these different visual mechanisms. We experimentally parasitized nests of the Common Redstart Phoenicurus phoenicurus, a regular host of a mimetic-egg laying Common Cuckoo Cuculus canorus host-race, using painted, immaculate 3D-printed model eggs in two geographically distant areas (Finland and Czech Republic). We found that directional chromatic contrasts better explained rejection behaviors in both parasitized (Finland) and non-parasitized (Czech Republic) host populations, as hosts rejected eggs that were noticeably browner, but not eggs that were noticeably bluer, than redstart eggs. These results support the paradigm of a single rejection threshold predicted by the directional chromatic contrast model and contribute to a growing generality of these patterns across diverse avian host-brood parasite systems.

Anti-parasitic egg rejection by great reed warblers (Acrocephalus arundinaceus) tracks differences along an eggshell color gradient.

One of the most effective defenses against avian brood parasitism is the rejection of the foreign egg from the host's nest. Until recently, most studies have tested whether hosts discriminate between own and foreign eggs based on the absolute differences in avian-perceivable eggshell coloration and maculation. However, recent studies suggest that hosts may instead contrast egg appearances across a directional eggshell color gradient. We assessed which discrimination rule best explained egg rejection by great reed warblers Acrocephalus arundinaceus, a frequent host to an egg-mimetic race of common cuckoos Cuculus canorus. We deployed 3D-printed model eggs varying in blue-green to brown coloration and in the presence of maculation. Using visual modeling, we calculated the absolute chromatic and achromatic just-noticeable differences (JNDs), as well as directional JNDs across a blue-green to brown egg color gradient, between host and model eggs. While most model eggs were rejected by great reed warblers, browner eggs were rejected with higher probability than more blue-green eggs, and the rejection probability did not depend on maculation. Directional egg color discrimination shown here and in a suite of recent studies on other host species may shape the cognitive decision rules that hosts use to recognize foreign eggs and affect the course of evolution in parasitic egg mimicry.

Seasonally sympatric but allochronic: differential expression of hypothalamic genes in a songbird during gonadal development.

Allochrony, the mismatch of reproductive schedules, is one mechanism that can mediate sympatric speciation and diversification. In songbirds, the transition into breeding condition and gonadal growth is regulated by the hypothalamic-pituitary-gonadal (HPG) axis at multiple levels. We investigated whether the difference in reproductive timing between two seasonally sympatric subspecies of dark-eyed juncos (Junco hyemalis) was related to gene expression along the HPG axis. During the sympatric pre-breeding stage, we measured hypothalamic and testicular mRNA expression of candidate genes via qPCR in captive male juncos. For hypothalamic mRNA, we found our earlier breeding subspecies had increased expression of gonadotropin-releasing hormone (GnRH) and decreased expression of androgen receptor, oestrogen receptor alpha and mineralocorticoid receptor (MR). Subspecies did not differ in expression of hypothalamic gonadotropin-inhibitory hormone (GnIH) and glucocorticoid receptor (GR). While our earlier breeding subspecies had higher mRNA expression of testicular GR, subspecies did not differ in testicular luteinizing hormone receptor, follicle-stimulating hormone receptor or MR mRNA expression levels. Our findings indicate increased GnRH production and decreased hypothalamic sensitivity to sex steroid negative feedback as factors promoting differences in the timing of gonadal recrudescence between recently diverged populations. Differential gene expression along the HPG axis may facilitate species diversification under seasonal sympatry.

Host defences against avian brood parasitism: an endocrine perspective.

Host defences against avian brood parasites are the outcome of well-documented coevolutionary arms races, yet important questions about variation in hosts' antiparasitic response traits remain poorly understood. Why are certain defences employed by some species or individuals and not by others? Here, we propose that understanding variability in and the evolution of host defences can be facilitated by the study of the underlying physiological mechanisms. Specifically, because antiparasitic strategies involve behaviours that have been shown to be hormonally regulated in other contexts, we hypothesize that host responses to brood parasites are likely to be mediated by related endocrine mechanisms. We outline the hallmarks of the endocrine bases of parasite defence-related avian behaviours, review the current understanding of antiparasitic host tactics and propose testable hypotheses about the hormonal mechanisms that may mediate host defences. We consider these mechanisms in a life-history framework and discuss how endocrine factors may shape variation in host defences. By providing a hypothesis-driven mechanistic framework for defences against parasitism, this perspective should stimulate the study of their endocrine bases to enhance our understanding of the intricate arms races in avian host-parasite systems.

Differential gene regulation underlies variation in melanic plumage coloration in the dark-eyed junco (Junco hyemalis).

Colour plays a prominent role in species recognition; therefore, understanding the proximate basis of pigmentation can provide insight into reproductive isolation and speciation. Colour differences between taxa may be the result of regulatory differences or be caused by mutations in coding regions of the expressed genes. To investigate these two alternatives, we studied the pigment composition and the genetic basis of coloration in two divergent dark-eyed junco (Junco hyemalis) subspecies, the slate-coloured and Oregon juncos, which have evolved marked differences in plumage coloration since the Last Glacial Maximum. We used HPLC and light microscopy to investigate pigment composition and deposition in feathers from four body areas. We then used RNA-seq to compare the relative roles of differential gene expression in developing feathers and sequence divergence in transcribed loci under common-garden conditions. Junco feathers differed in eumelanin and pheomelanin content and distribution. Within subspecies, in lighter feathers melanin synthesis genes were downregulated (including PMEL, TYR, TYRP1, OCA2 and MLANA), and ASIP was upregulated. Feathers from different body regions also showed differential expression of HOX and WNT genes. Feathers from the same body regions that differed in colour between the two subspecies showed differential expression of ASIP and three other genes (MFSD12, KCNJ13 and HAND2) associated with pigmentation in other taxa. Sequence variation in the expressed genes was not related to colour differences. Our findings support the hypothesis that differential regulation of a few genes can account for marked differences in coloration, a mechanism that may facilitate the rapid phenotypic diversification of juncos.

The effect of chronic and acute stressors, and their interaction, on testes function: an experimental test during testicular recrudescence.

Organisms are expected to invest less in reproduction in response to a stressor, but theory predicts that this effect should depend on the frequency and duration of stressors in the environment. Here, we investigated how an acute stressor affected testes function in a songbird, and how chronic stressors influenced the acute stress response. We exposed male dark-eyed juncos (Junco hyemalis) either to chronic or minimal (control) disturbance during testicular recrudescence, after which we measured baseline testosterone, testosterone after an acute handling stressor, and capacity to produce testosterone after hormonal stimulation. In a 2×2 design, we then killed males from the two chronic treatment groups either immediately or after an acute stressor to investigate the effect of long- and short-term stressors on the testicular transcriptome. We found that chronically disturbed birds had marginally lower baseline testosterone. The acute stressor suppressed testosterone in control birds, but not in the chronic disturbance group. The ability to elevate testosterone did not differ between the chronic treatments. Surprisingly, chronic disturbance had a weak effect on the testicular transcriptome, and did not affect the transcriptomic response to the acute stressor. The acute stressor, on the other hand, upregulated the cellular stress response and affected expression of genes associated with hormonal stress response. Overall, we show that testicular function is sensitive to acute stressors but surprisingly robust to long-term stressors, and that chronic disturbance attenuates the decrease in testosterone in response to an acute stressor.

Chronological and Biological Age Predict Seasonal Reproductive Timing: An Investigation of Clutch Initiation and Telomeres in Birds of Known Age.

Female vertebrates that breed earlier in the season generally have greater reproductive success. However, evidence suggests that breeding early may be costly, thus leading to the prediction that females with fewer future reproductive events will breed earlier in the season. While chronological age is a good indicator of remaining life span, telomere lengths may also be good biomarkers of longevity as they potentially reflect lifetime wear and tear (i.e., biological age). We examined whether variation in the timing of the first seasonal clutch was related to age and telomere length in female dark-eyed juncos (Junco hyemalis), predicting that older females and those with shorter telomeres would breed earlier. Both predictions held true and were independent of each other, as telomere length did not significantly vary with age. These results suggest that females may adjust their reproductive effort based on both chronological and biological age.

Effect of acute stressor on reproductive behavior differs between urban and rural birds.

The life-history trade-off between self-maintenance and reproduction posits that investment in one function decreases investment in the other. Manipulating the costs and benefits of functions involved in a trade-off may alter this interaction. Here we ask whether investment in self-maintenance during a stress response alters territorial behavior in wild Dark-eyed Juncos and whether rural and urban birds, which are known to differ in the magnitude of the stress response (greater in rural), also differ in the degree to which stress reduces territorial behavior. In rural and urban habitats, we measured territorial behavior using song playback, followed by either an acute stressor (capture and collection of a blood sample) or a nonstressful control situation. The following day, we again measured territorial behavior, predicting greater reduction in territorial behavior in individuals exposed to the stressor but a lesser reduction in territorial behavior in the urban as compared to the rural environment. We further assessed individual and population differences in response to stressors by measuring flight initiation distance, breath rate, and corticosterone levels in the blood. The rural population had a higher physiological and behavioral stress response than the urban population, and acute capture stress had a lasting (24 h) negative effect on territorial behavior, but only in the rural habitat. However, individual-level differences in measures of the stress response did not explain variation in the impact of stress on territorial behavior. Our findings show that stressors can have a negative effect on territorial behavior, but that this effect may differ between populations that vary in their stress ecology.

The excuse principle can maintain cooperation through forgivable defection in the Prisoner's Dilemma game.

Reciprocal altruism describes a situation in which an organism acts in a manner that temporarily reduces its fitness while increasing another organism's fitness, but there is an ultimate fitness benefit based on an expectation that the other organism will act in a similar manner at a later time. It creates the obvious dilemma in which there is always a short-term benefit to cheating, therefore cooperating individuals must avoid being exploited by non-cooperating cheaters. This is achieved by following various decision rules, usually variants of the tit-for-tat (TFT) strategy. The strength of TFT, however, is also its weakness-mistakes in implementation or interpretation of moves, or the inability to cooperate, lead to a permanent breakdown in cooperation. We show that pied flycatchers (Ficedula hypoleuca) use a TFT with an embedded 'excuse principle' to forgive the neighbours that were perceived as unable to cooperate during mobbing of predators. The excuse principle dramatically increases the stability of TFT-like behavioural strategies within the Prisoner's Dilemma game.

Variation in candidate genes CLOCK and ADCYAP1 does not consistently predict differences in migratory behavior in the songbird genus Junco.

Recent studies exploring the molecular genetic basis for migratory variation in animals have identified polymorphisms in two genes ( CLOCK and ADCYAP1) that are linked to circadian rhythms and correlate with migratory propensity and phenology among individuals and populations. Results from these initial studies are mixed, however, and additional data are needed to assess the generality and diversity of the molecular mechanisms that regulate the biology of migration. We sequenced CLOCK and ADCYAP1 in 15 populations across the two species of the avian genus Junco, a North American lineage in which multiple recently diverged subspecies and populations range from sedentary to long-distance migrants. We found no consistent associations between allele length and migratory status across the genus for either CLOCK or ADCYAP1. However, within two subspecies groups, populations that migrate longer distances have longer CLOCK alleles on average. Additionally, there was a positive relationship between ADCYAP1 allele length and migratory restlessness (zugunruhe) among individuals within one of two captive populations studied-a result similar to those reported previously within captive blackcaps ( Sylvia atricapilla). We conclude that, while both ADCYAP1 and CLOCK may correlate with migratory propensity within or among certain populations or species, previously identified relationships between migratory behavior and sequence variants cannot be easily generalized across taxa.