Parental age does not influence offspring telomeres during early life in common gulls (Larus canus).

Parental age can affect offspring telomere length through heritable and epigenetic-like effects, but at what stage during development these effects are established is not well known. To address this, we conducted a cross-fostering experiment in common gulls (Larus canus) that enabled us distinguish between pre- and post-natal parental age effects on offspring telomere length. Whole clutches were exchanged after clutch completion within and between parental age classes (young and old) and blood samples were collected from chicks at hatching and during the fastest growth phase (11 days later) to measure telomeres. Neither the ages of the natal nor the foster parents predicted the telomere length or the change in telomere lengths of their chicks. Telomere length (TL) was repeatable within chicks, but increased across development (repeatability = 0.55, intraclass correlation coefficient within sampling events 0.934). Telomere length and the change in telomere length were not predicted by post-natal growth rate. Taken together, these findings suggest that in common gulls, telomere length during early life is not influenced by parental age or growth rate, which may indicate that protective mechanisms buffer telomeres from external conditions during development in this relatively long-lived species.

Corticosterone levels correlate in wild-grown and lab-grown feathers in greenfinches (Carduelis chloris) and predict behaviour and survival in captivity.

Level of corticosterone (CORT), which is a predominant glucocorticoid in birds, has become the main indicator for assessing the stress level of birds in ecological studies. Feather corticosterone (CORTf) provides information about corticosterone levels during feather growth, however, the underlying causes of individual variation of CORTf between individuals and individual persistency of CORTf are not yet fully understood. Therefore, this study addresses individual consistency in CORTf and the association of variation in CORTf with behaviour that results in damage to tail feathers. We studied relations between CORTf, plasma CORT, and behaviour in wild-caught male greenfinches in captivity. CORTf in wild-grown feathers correlated positively with CORTf in lab-grown feathers. CORTf levels were about 20% lower in lab-grown feathers than in those grown in the wild. Four birds that died in captivity had significantly higher average CORTf levels in wild-grown feathers than the survivors. Plasma CORT levels of two measurements taken in the lab seven days apart correlated positively, however, no correlations between plasma CORT and CORTf were found. In order to study the link between CORTf and behaviour, the extent of tail damage from flapping against cage bar was assessed. Contrary to our prediction, birds with higher CORTf had less tail damage. This study adds to the evidence that CORTf levels can be considered as informative markers of some persistent component of individual phenotypic quality that can predict survival under standardized laboratory conditions.

Uropygial gland size: a marker of phenotypic quality that shows no senescence in a long-lived seabird.

Studies of senescence in the wild have traditionally focused on traits like survival or fecundity. Although efforts to measure other salient phenotypic traits and markers of relevant physiological processes are rapidly increasing, traits related to self-maintenance remain understudied in the context of aging. Uropygial or preen gland is a holocrine gland, exclusive to birds, directly linked to self-maintenance of the quality of plumage. We measured the size of uropygial glands of common gulls (Larus canus) in a cross sectional manner in order to test whether it shows the similar age-related decline as reproductive traits previously recorded in this species. Gulls with larger glands started breeding earlier in the season, indicating that gland size is a marker of individual phenotypic quality. We found a senescent decline in the onset of breeding and the size of white wing patches, a sexually dimorphic ornamental trait, while in contrast, preen gland increased with advancing age. This finding supports the view of life-history theory that in long-lived species whose lifetime reproductive success depends heavily on lifespan, self-maintenance is prioritized over reproduction. Altogether our results support the concept that senescence in the wild can be asynchronous for traits related to maintenance versus reproduction.

Morphometric traits predict educational attainment independently of socioeconomic background.

BACKGROUND: Aim of this study is to describe the relationship between anthropometric traits and educational attainment among Estonian schoolchildren born between 1937 and 1962. We asked whether height, cranial volume and face width (a testosterone-dependent trait), measured in childhood predict later educational attainment independently of each other, family socioeconomic position (SEP) and sex. Associations between morphometric traits and education and their interactions with biosocial variables are of scholarly importance because higher education is nearly universally associated with low fertility in women, and often with high fertility in men. Hence, morphometric traits associated with educational attainment are targeted by natural selection and describing the exact nature of these associations is relevant for understanding the current patterns of evolution of human body size. METHODS: Data on morphometric measurements and family background of 11,032 Estonian schoolchildren measured between seven and 19 years of age were obtained from the study performed by Juhan Aul between 1956 and 1969. Ordinal logistic regression was used for testing the effects of morphometric traits, biosocial variables and their interaction on the cumulative probability of obtaining education beyond primary level. RESULTS: Of biosocial variables, family SEP was the most important determinant of educational attainment, followed by the sex, rural vs urban origin and the number of siblings. No significant interactions with morphometric traits were detected, i.e., within each category of SEP, rural vs urban origin and sex, taller children and those with larger heads and relatively narrower faces were more likely to proceed to secondary and/or tertiary education. The effect of height on education was independent of cranial volume, indicating that taller children did not obtain more educations because their brains were larger than those of shorter children; height per se was important. CONCLUSIONS: Our main finding - that adjusting for other morphometric traits and biosocial variables, morphometric traits still robustly predicted educational attainment, is relevant for understanding the current patterns of evolution of human body size. Our findings suggest that fecundity selection acting on educational attainment could be partly responsible for the concurrent selection for smaller stature and cranial volume in women and opposite trends in men.

Age-specific patterns of maternal investment in common gull egg yolk.

While the general patterns of age-specific changes in reproductive success are quite well established in long-lived animals, we still do not know if allocation patterns of maternally transmitted compounds are related to maternal age. We measured the levels of yolk testosterone, carotenoids and vitamins A and E in a population of known-aged common gulls (Larus canus) and found an age-specific pattern in yolk lutein and vitamin A concentrations. Middle-aged mothers allocated more of these substances to yolk compared to young and old mothers. These results can be explained through differences in age-specific foraging, absorption or deposition patterns of carotenoids and vitamins into yolk. If these molecules play a role in antioxidant defence and immune modulation, our results suggest a possible physiological pathway underlying the age-specific changes in reproductive success of long-lived birds in the wild.

Viability selection affects black but not yellow plumage colour in greenfinches.

Much of the debate surrounding the selective forces responsible for the expression of conspicuous plumage colouration is centred on the question of precisely which individual qualities are signalled by carotenoid- and melanin-based pigments. To examine this and other related issues, we performed viability selection analyses in wild-caught captive male greenfinches (Carduelis chloris) in Estonia during winters between 2003 and 2014. Based on our measurements, birds with a darker black eumelanin-based colouration of tail feathers survived better than those whose tail feathers had a paler black colouration. The carotenoid-based yellow colouration of the same feathers was not associated with mortality in captivity and showed much less between-year variation in the field than the black colouration. Between year-variation in the black (but not yellow) colouration of feathers was parallel in wild-grown feathers (on birds in the wild) and laboratory-grown ones (on birds held temporarily in captivity). Taken together, these findings imply that eumelanotic colouration in greenfinches is currently under selection and suggest the presence of sufficient genetic variation for a rapid response to selection. In particular, tail feathers have become darker black since the emergence of avian trichomonosis, which is known to selectively kill paler individuals.

Skin pentosidine and telomere length do not covary with age in a long-lived seabird.

The questions about why and how senescence occurs in the wild are among the most pertinent ones in evolutionary ecology. Telomere length is a commonly used marker for aging, while other biomarkers of aging have received considerably less attention. Here we studied how another potent indicator of aging-skin pentosidine concentration-relates to age and blood telomere length in a long-lived seabird with well-documented reproductive senescence. We found no associations between telomere length, skin pentosidine and chronological age in male common gulls (Larus canus), aging from 2 to 30 years. However, the variance in telomere length was 4.6 times higher among the birds older than 13 years, which hints at relaxed selection on telomere length among the birds that have passed their prime age of reproduction. These results suggest that physiological and chronological ages may be largely uncoupled in our study system. Furthermore, our findings do not support a hypothesis about the presence of a common physiological factor (e.g., such as oxidative stress) that would cause covariation between two independent markers of aging.

Father's death does not affect growth and maturation but hinders reproduction: evidence from adolescent girls in post-war Estonia.

The popular concept of predictive-adaptive responses poses that girls growing up without a father present in the family mature and start reproduction earlier because the father's absence is a cue for environmental harshness and uncertainty that favours switching to a precocious life-history strategy. Most studies supporting this concept have been performed in situations where the father's absence is caused by divorce or abandonment. Using a dataset of Estonian adolescent girls who had lost their fathers over the period of World War II, we show that father's death did not affect the rate of pubertal maturation (assessed on the basis of development of breasts and axillary hair) or growth. Father's death did not affect the age of first birth but, contrary to predictions, reduced lifetime reproductive success. Our findings thus do not support the concept of predictive-adaptive responses and suggest that alternative explanations for covariation between fatherlessness and early maturation are required.

Profile of whole blood gene expression following immune stimulation in a wild passerine.

BACKGROUND: Immunoecology aims to explain variation among hosts in the strength and efficacy of immunological defences in natural populations. This requires development of biomarkers of the activation of the immune system so that they can be collected non-lethally and sampled from small amounts of easily obtainable tissue. We used transcriptome profiling in wild greenfinches (Carduelis chloris) to detect whole blood transcripts that most profoundly indicate upregulation of antimicrobial defences during acute phase response. The more general aim of this study was to obtain a functional annotation of a substantial portion of the greenfinch transcriptome that would enable to gain access to more specific genomic tools in subsequent studies. The birds received either bacterial lipopolysaccharide or saline injections and RNA-seq transcriptional profiling was performed 12 h after treatment to provide initial functional annotation of the transcriptome and assess whole blood response to immune stimulation. RESULTS: A total of 66,084 transcripts were obtained from de novo Trinty assembly, out of which 23,153 could be functionally annotated. Only 1,911 of these were significantly upregulated or downregulated. The manipulation caused marked upregulation of several transcripts related to immune activation. These included avian-specific antimicrobial agents avidin and gallinacin, but also some more general host response genes, such as serum amyloid A protein, lymphocyte antigen 75 and copper-transporting ATPase 1. However, links with avian immunity for most differentially regulated transcripts remained rather hypothetical, as a large set of differentially expressed transcripts lacked functional annotation. CONCLUSIONS: This appears to be the first large scale transcriptional profiling of immune function in passerine birds. The transcriptomic data obtained suggest novel markers for the assessment of the immunological state of wild passerines. Characterizing the function of those possible novel infection markers would assist future vertebrate genome annotation. The extensive sequence information collected enables to identify possible target and housekeeping genes needed to gain access to more specific genomic tools in future studies.

High feather corticosterone indicates better coccidian infection resistance in greenfinches.

Differential exposure or sensitivity to stressors can have substantial effects on the variation in immune responsiveness of animals. However, the questions about the causes and consequences of these processes have remained largely unclear, particularly as regards wild animals and their natural pathogens. Here we ask how a potential marker of stress responses, the feather corticosterone (CORT) content, reflects the resistance to an experimental infection with natural coccidian parasites in wild-caught captive greenfinches (Carduelis chloris). CORT content of tail feathers grown in captivity correlated positively with a behavioural measure of captivity-intolerance, i.e., the amount of damage accrued to tail feathers in captivity that results from flapping against cage bars. This finding is consistent with an idea that feather CORT reflects the amount of stress experienced during feather growth. Experimental infection with heterologous coccidian strains increased feather CORT levels. Birds with highest feather CORT levels appeared most resistant to new infection, assessed on the basis of parasite oocyst shedding at the peak phase of infection. Birds with highest feather CORT levels also cleared the infection faster than the birds with lower feather CORT levels. These findings provide the first evidence about positive covariation between feather CORT and resistance to a natural pathogen in a wild bird species. Assuming that feather CORT levels reflect circulating hormone titres, these findings suggest that parasite-mediated selection may contribute to maintenance of phenotypes with high corticosterone responsiveness to stress, despite potential negative behavioural consequences.

Survival costs and benefits of reproduction: A register-based study in 20th century Estonia.

Patterns of individual variation in lifespan and senescence depend on the associations between parental survival and reproductive rates. We studied the associations between parity and survival among 579,271 Estonians born between 1905 and 1945 and in a cohort with a completed lifespan born in 1905-1927. For this cohort, selection for increased lifespan operated on both sexes, but it was stronger in men than in women. However, the median lifespan increased between the subsequent cohorts in women but stagnated in men. Selection for longer lifespan was caused by the below-average lifespan of individuals with no or single offspring. Despite a general positive selection for lifespan, survival costs of reproduction were also detected among a relatively small proportion of individuals with high parities, as mothers of two and fathers of two and three children had the highest median lifespans. Fathers of more than six children had better survival than fathers of few children in their reproductive age, but this association reversed after age 70. The reversal of this association between survival and parity at old age indicates that relative mortality risks between those with lower versus higher parities change across ages, as predicted by the antagonistic pleiotropy theory of aging.

On the methodological limitations of detecting oxidative stress: effects of paraquat on measures of oxidative status in greenfinches.

Oxidative stress (OS) is widely believed to be responsible for the generation of trade-offs in evolutionary ecology by means of constraining investment into a number of components of fitness. Yet, progress in understanding the true role of OS in ecology and evolution has remained elusive. Interpretation of current findings is particularly hampered by the scarcity of experiments demonstrating which of the many available parameters of oxidative status respond most sensitively to and are relevant for measuring OS. We addressed these questions in wild-caught captive greenfinches (Carduelis chloris) by experimental induction of OS by administration of the pro-oxidant compound paraquat with drinking water. Treatment induced 50% mortality, a significant drop in body mass and an increase in oxidative DNA damage and glutathione levels in erythrocytes among the survivors of the high paraquat (0.2 g l(-1) over 7 days) group. Samples taken 3 days after the end of paraquat treatment showed no effect on the peroxidation of lipids (plasma malondialdehyde), carbonylation of proteins (in erythrocytes), parameters of plasma antioxidant protection (total antioxidant capacity and oxygen radical absorbance), uric acid or carotenoids. Our findings of an increase in one marker of damage and one marker of protection from the multitude of measured variables indicate that detection of OS is difficult even under the most stringent experimental induction of oxidative insult. We hope that this study highlights the need for reconsideration of over-simplistic models of OS and draws attention to the limitations of detection of OS due to time-lagged and hormetic upregulation of protective mechanisms. This study also underpins the diagnostic value of measurement of oxidative damage to DNA bases and assessment of erythrocyte glutathione levels.

Dexamethasone inhibits corticosterone deposition in feathers of greenfinches.

Corticosterone (CORT) content of feathers is a potent source of information about activation of hypothalamus-pituitary-adrenal (HPA) axis during feather growth, which is used for assessment of well-being and stress history of individuals and populations in avian studies. However, little is known about factors affecting deposition of CORT into feathers and how feather CORT covaries with other markers of stress imposed upon individuals during feather growth. We addressed these questions by measuring CORT levels in feathers of wild-caught greenfinches (Carduelis chloris) brought into captivity. One tail feather was removed from all the birds upon arrival to the laboratory and the CORT levels of replacement feathers, grown in captivity were recorded. The birds were subjected to treatments of immune activation (by injection of phytohaemagglutinin) and synthetic glucocorticoid (dexamethasone, DEX) administration. Only DEX injection affected feather CORT levels. DEX-injected birds deposited on average 37% less of CORT in their feathers than saline-injected birds. Despite significant effects of DEX and immune activation treatments on differential leukocyte counts, we did not find any correlations between CORT and leukocyte hemoconcentrations or heterophil/lymphocyte ratios (a haematological index of stress), measured at three stages of feather growth. Our findings provide novel evidence that feather CORT levels are sensitive to manipulation of hormonal balance of birds, thereby supporting the diagnostic value of feather CORT measurements. However, we did not find any evidence about covariation between feather CORT and other markers of stress perceived during the period of feather growth. This calls for further research on information content of feather CORT, preferably in experiments manipulating more diverse array of psychological, immunological and abiotic stressors.

Mothers of small-bodied children and fathers of vigorous sons live longer.

Life-history traits (traits directly related to survival and reproduction) co-evolve and materialize through physiology and behavior. Accordingly, lifespan can be hypothesized as a potentially informative marker of life-history speed that subsumes the impact of diverse morphometric and behavioral traits. We examined associations between parental longevity and various anthropometric traits in a sample of 4,000-11,000 Estonian children in the middle of the 20th century. The offspring phenotype was used as a proxy measure of parental genotype, so that covariation between offspring traits and parental longevity (defined as belonging to the 90th percentile of lifespan) could be used to characterize the aggregation between longevity and anthropometric traits. We predicted that larger linear dimensions of offspring associate with increased parental longevity and that testosterone-dependent traits associate with reduced paternal longevity. Twelve of 16 offspring traits were associated with mothers' longevity, while three traits (rate of sexual maturation of daughters and grip strength and lung capacity of sons) robustly predicted fathers' longevity. Contrary to predictions, mothers of children with small bodily dimensions lived longer, and paternal longevity was not linearly associated with their children's body size (or testosterone-related traits). Our study thus failed to find evidence that high somatic investment into brain and body growth clusters with a long lifespan across generations, and/or that such associations can be detected on the basis of inter-generational phenotypic correlations.

Heterophil/lymphocyte ratios predict the magnitude of humoral immune response to a novel antigen in great tits (Parus major).

Animals display remarkable individual variation in their capacity to mount immune responses against novel antigens. According to the life-history theory, this variation is caused by the costs of immune responses to the hosts. We studied one of such potential costs, depletion of somatic resources in wintering wild-caught captive passerines, the great tits (Parus major) by immune challenging the birds with a novel antigen, killed Brucella abortus (BA) suspension. We found that despite mild temperature conditions in captivity and ad libitum availability of food, immune challenge depleted somatic resources (as indicated by a body mass loss) and elevated relative proportion of heterophils to lymphocytes (H/L ratio) in the peripheral blood of birds. However, body mass loss did not covary with an increase in H/L ratios between two sampling events, which indicates that these two markers of health state describe different aspects of individual physiological condition. Antibody titres were not associated with the extent of body mass loss during the development of immune response, which shows that the somatic cost of immune response was not proportional to the amount of antibody produced. Birds with high pre-immunisation H/L ratios mounted weaker antibody response, which is indicative of stress-induced suppression of humoral immune response and is consistent with the concept of an antagonistic cross-regulation between different components of the immune system. The latter finding suggests a novel diagnostic value of H/L ratios, which reinforces the utility of this simple haematological index for prediction of the outcomes of complicated immune processes.

Sexually antagonistic selection on educational attainment and body size in Estonian children.

Natural selection is a key mechanism of evolution, which results from the differential reproduction of phenotypes. We describe fecundity selection at different parity transitions on 15 anthropometric traits and educational attainment in Estonian children sampled in the middle of 20th century. The direction of selection on educational attainment and bodily traits was sexually antagonistic, and it occurred via different parity transitions in boys and girls. Compared to boys with primary education, obtaining tertiary education was associated with 3.5 times and secondary education two times higher odds of becoming a father. Transition to motherhood was not related to educational attainment, while education above primary was associated with lower odds (OR = 0.5-0.7) to progression to parities above one and two. Selection on anthropometric traits occurred almost exclusively via childlessness in boys, while among the girls, most of the traits that were associated with becoming a mother were additionally associated with a transition from one child to higher parities. Male (but not female) fitness was thus primarily determined by traits related to mating success. Selection favored stronger and larger boys and smaller girls. Selection on girls favored some traits that associate with perceived femininity, while other feminine traits were selected against.

Behavioural trait covaries with immune responsiveness in a wild passerine.

Immune system is highly integrated with the nervous and endocrine systems, which is thought to result in covariation between behavioural syndromes and stress- and immune-associated diseases. Very little is known about the associations between behaviour and immune traits in wild animals. Here we describe such an association in passerine birds, the greenfinches (Carduelis chloris). When wild-caught greenfinches are brought into captivity, some individuals damage their tail feathers against cage walls due to excited behaviour, while others retain their feathers in intact condition. We show that damage to tail feathers was associated with flapping flight movements and the frequency of such flapping bouts was individually consistent over 57 days. Birds with intact tails, i.e., relatively 'calm' individuals mounted stronger antibody response to a novel Brucella abortus antigen and their circulating phagocytes were capable of producing stronger oxidative burst in response to stimulation with bacterial lipopolysaccharide in vitro. As the behavioural trait was assessed 13-25 days before measuring immune responsiveness, our results demonstrate that individuals' coping styles with captivity predicted how these individuals would respond to forthcoming immune challenges. This is a novel evidence about covariation between immune responsiveness and a behavioural trait in a wild-caught animal.

Carotenoid intake does not affect immune-stimulated oxidative burst in greenfinches.

Carotenoid-based integument colouration is extremely widespread in the animal kingdom. It has been hypothesized that carotenoid colouration is used for communicating the health status of the bearers because carotenoids are efficient immunomodulators or antioxidants. However, the latter argument has been recently debated and the mechanisms by which carotenoids modulate immunity or oxidative balance are poorly known. We performed an experiment on wild-caught captive greenfinches, passerine birds with carotenoid-based plumage colouration, in order to test whether dietary carotenoid supplementation affects immune-stimulated oxidative burst of phagocytes in the whole blood and humoral immune response to a novel antigen, Brucella abortus (BA). Additionally, we tested whether immune stimulation with bacterial lipopolysaccharide (LPS) affects blood carotenoid levels. We thus tested the effects of carotenoids on the oxidative burst of phagocytes under neutral conditions and during in vivo immune challenge. LPS injection depleted plasma carotenoids, indicating involvement of these phytochemicals in the immune response. However, we did not find any evidence that manipulation of carotenoid intake had modulated anti-BA antibody production, LPS-stimulated oxidative burst of phagocytes, or basal levels of circulating reactive oxygen species. This indicates that carotenoid intake does not affect endogenous production of reactive oxygen species by immune cells. This finding is consistent with the view that carotenoids are unlikely to provide a direct link between oxidative stress and colouration. However, it remains to be tested whether the oxidative burst of phagocytes induced in our experiment actually inflicts oxidative damage and whether carotenoids play a role in the attenuation of such potential damages.

Effects of carotenoids, immune activation and immune suppression on the intensity of chronic coccidiosis in greenfinches.

Allocation trade-offs of carotenoids between their use in the immune system and production of integumentary colouration have been suggested as a proximate mechanism maintaining honesty of signal traits. We tested how dietary carotenoid supplementation, immune activation and immune suppression affect intensity of coccidian infection in captive greenfinches Carduelis chloris, a passerine with carotenoid-based plumage. Immune activation with phytohaemagglutinin (PHA) decreased body mass among birds not supplemented with lutein, while among the carotenoid-fed birds, PHA had no effect on mass dynamics. Immune suppression with dexamethasone (DEX) induced loss of body mass and reduced the swelling response to PHA. DEX and PHA increased the concentration of circulating heterophils. Lutein supplementation increased plasma carotenoid levels but had no effect on the swelling response induced by PHA. PHA and DEX treatments did not affect plasma carotenoids. Immune stimulation by PHA suppressed the infection, but only among carotenoid-supplemented birds. Priming of the immune system can thus aid in suppressing chronic infection but only when sufficient amount of carotenoids is available. Our experiment shows the importance of carotenoids in immune response, but also the complicated nature of this impact, which could be the reason for inconsistent results in studies investigating the immunomodulatory effects of carotenoids. The findings about involvement of carotenoids in modulation of an immune response against coccidiosis suggest that carotenoid-based ornaments may honestly signal individuals' ability to manage chronic infections.

Effects of chronic leptin administration on nitric oxide production and immune responsiveness of greenfinches.

Leptin and nitric oxide (NO) are both important messengers in intra- and intercellular communication systems in vertebrates. Several studies have demonstrated an involvement of both substances in the immune response. Here we tested the effects of chronic leptin and anti-leptin treatments on the NO production and phytohaemagglutinin- (PHA) induced cutaneous inflammatory response in a wild passerine, the greenfinch (Carduelis chloris). Plasma leptin levels of individual birds were consistent in time but could be still temporarily increased by administration of recombinant chicken leptin. Increase of plasma leptin was also induced by administration of anti-leptin, which can be most likely explained by increased endogenous leptin production due to disruption of signalling pathways. Contrary to previous findings in mammals, leptin administration reduced systemic NO production. Leptin increased cutaneous swelling response to PHA. This immune-enhancing effect was observable despite the similar plasma leptin levels of leptin-treated and control birds at the time of measurement of immune responses, i.e., 9 days after start of the treatments. This provides evidence for a delayed or long-term potentiation of the cells and cytokines involved. The effects of leptin administration on NO production and immune responsiveness were age-dependent, which indicates the complexity of underlying regulatory mechanisms.