From maternal glucocorticoid and thyroid hormones to epigenetic regulation of offspring gene expression: An experimental study in a wild bird species.

Offspring phenotype at birth is determined by its genotype and the prenatal environment including exposure to maternal hormones. Variation in both maternal glucocorticoids and thyroid hormones can affect offspring phenotype, but the underlying molecular mechanisms, especially those contributing to long-lasting effects, remain unclear. Epigenetic changes (such as DNA methylation) have been postulated as mediators of long-lasting effects of early-life environment. In this study, we determined the effects of elevated prenatal glucocorticoid and thyroid hormones on handling stress response (breath rate) as well as DNA methylation and gene expression of glucocorticoid receptor (GR) and thyroid hormone receptor (THR) in great tits (Parus major). Eggs were injected before incubation onset with corticosterone (the main avian glucocorticoid) and/or thyroid hormones (thyroxine and triiodothyronine) to simulate variation in maternal hormone deposition. Breath rate during handling and gene expression of GR and THR were evaluated 14 days after hatching. Methylation status of GR and THR genes was analyzed from the longitudinal blood cells sampled 7 and 14 days after hatching, as well as the following autumn. Elevated prenatal corticosterone level significantly increased the breath rate during handling, indicating an enhanced metabolic stress response. Prenatal corticosterone manipulation had CpG-site-specific effects on DNA methylation at the GR putative promoter region, while it did not significantly affect GR gene expression. GR expression was negatively associated with earlier hatching date and chick size. THR methylation or expression did not exhibit any significant relationship with the hormonal treatments or the examined covariates, suggesting that TH signaling may be more robust due to its crucial role in development. This study provides some support to the hypothesis suggesting that maternal corticosterone may influence offspring metabolic stress response via epigenetic alterations, yet their possible adaptive role in optimizing offspring phenotype to the prevailing conditions, context-dependency, and the underlying molecular interplay needs further research.

Environmental Drivers of Growth and Oxidative Status during Early Life in a Long-Lived Antarctic Seabird, the Adélie Penguin.

AbstractIn vertebrates, developmental conditions can have long-term effects on individual performance. It is increasingly recognized that oxidative stress could be one physiological mechanism connecting early-life experience to adult phenotype. Accordingly, markers of oxidative status could be useful for assessing the developmental constraints encountered by offspring. Although some studies have demonstrated that developmental constraints are associated with high levels of oxidative stress in offspring, it remains unclear how growth, parental behavior, and brood competition may altogether affect oxidative stress in long-lived species in the wild. Here, we investigated this question in a long-lived Antarctic bird species by testing the impact of brood competition (e.g., brood size and hatching order) on body mass and on two markers of oxidative damage in Adélie penguin chicks. We also examined the influence of parental effort (i.e., foraging trip duration) and parental body condition on chick body mass and oxidative damage. First, we found that brood competition and parental traits had significant impacts on chick body mass. Second, we found that chick age and, to a lesser extent, chick body mass were two strong determinants of the levels of oxidative damage in Adélie penguin chicks. Finally, and importantly, we also found that brood competition significantly increased the levels of one marker of oxidative damage and was associated with a lower survival probability. However, parental effort and parental condition were not significantly linked to chick levels of oxidative damage. Overall, our study demonstrates that sibling competition can generate an oxidative cost even for this long-lived Antarctic species with a limited brood size (maximum of two chicks).

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.

The role of individual variability on the predictive performance of machine learning applied to large bio-logging datasets.

Animal-borne tagging (bio-logging) generates large and complex datasets. In particular, accelerometer tags, which provide information on behaviour and energy expenditure of wild animals, produce high-resolution multi-dimensional data, and can be challenging to analyse. We tested the performance of commonly used artificial intelligence tools on datasets of increasing volume and dimensionality. By collecting bio-logging data across several sampling seasons, datasets are inherently characterized by inter-individual variability. Such information should be considered when predicting behaviour. We integrated both unsupervised and supervised machine learning approaches to predict behaviours in two penguin species. The classified behaviours obtained from the unsupervised approach Expectation Maximisation were used to train the supervised approach Random Forest. We assessed agreement between the approaches, the performance of Random Forest on unknown data and the implications for the calculation of energy expenditure. Consideration of behavioural variability resulted in high agreement (> 80%) in behavioural classifications and minimal differences in energy expenditure estimates. However, some outliers with < 70% of agreement, highlighted how behaviours characterized by signal similarity are confused. We advise the broad bio-logging community, approaching these large datasets, to be cautious when upscaling predictions, as this might lead to less accurate estimates of behaviour and energy expenditure.

Effect of prenatal glucocorticoids and thyroid hormones on developmental plasticity of mitochondrial aerobic metabolism, growth and survival: an experimental test in wild great tits.

Developmental plasticity is partly mediated by transgenerational effects, including those mediated by the maternal endocrine system. Glucocorticoid and thyroid hormones may play central roles in developmental programming through their action on metabolism and growth. However, the mechanisms by which they affect growth and development remain understudied. One hypothesis is that maternal hormones directly affect the production and availability of energy-carrying molecules (e.g. ATP) by their action on mitochondrial function. To test this hypothesis, we experimentally increased glucocorticoid and thyroid hormones in wild great tit eggs (Parus major) to investigate their impact on offspring mitochondrial aerobic metabolism (measured in blood cells), and subsequent growth and survival. We show that prenatal glucocorticoid supplementation affected offspring cellular aerobic metabolism by decreasing mitochondrial density, maximal mitochondrial respiration and oxidative phosphorylation, while increasing the proportion of the maximum capacity being used under endogenous conditions. Prenatal glucocorticoid supplementation only had mild effects on offspring body mass, size and condition during the rearing period, but led to a sex-specific (females only) decrease in body mass a few months after fledging. Contrary to our expectations, thyroid hormone supplementation did not affect offspring growth or mitochondrial metabolism. Recapture probability as juveniles or adults was not significantly affected by prenatal hormonal treatment. Our results demonstrate that prenatal glucocorticoids can affect post-natal mitochondrial density and aerobic metabolism. The weak effects on growth and apparent survival suggest that nestlings were mostly able to compensate for the transient decrease in mitochondrial aerobic metabolism induced by prenatal glucocorticoids.

Acceleration predicts energy expenditure in a fat, flightless, diving bird.

Energy drives behaviour and life history decisions, yet it can be hard to measure at fine scales in free-moving animals. Accelerometry has proven a powerful tool to estimate energy expenditure, but requires calibration in the wild. This can be difficult in some environments, or for particular behaviours, and validations have produced equivocal results in some species, particularly air-breathing divers. It is, therefore, important to calibrate accelerometry across different behaviours to understand the most parsimonious way to estimate energy expenditure in free-living conditions. Here, we combine data from miniaturised acceleration loggers on 58 free-living Adélie penguins with doubly labelled water (DLW) measurements of their energy expenditure over several days. Across different behaviours, both in water and on land, dynamic body acceleration was a good predictor of independently measured DLW-derived energy expenditure (R2 = 0.72). The most parsimonious model suggested different calibration coefficients are required to predict behaviours on land versus foraging behaviour in water (R2 = 0.75). Our results show that accelerometry can be used to reliably estimate energy expenditure in penguins, and we provide calibration equations for estimating metabolic rate across several behaviours in the wild.

Born to be young? Prenatal thyroid hormones increase early-life telomere length in wild collared flycatchers.

The underlying mechanisms of the lifelong consequences of prenatal environmental condition on health and ageing remain little understood. Thyroid hormones (THs) are important regulators of embryogenesis, transferred from the mother to the embryo. Since prenatal THs can accelerate early-life development, we hypothesized that this might occur at the expense of resource allocation in somatic maintenance processes, leading to premature ageing. Therefore, we investigated the consequences of prenatal TH supplementation on potential hallmarks of ageing in a free-living avian model in which we previously demonstrated that experimentally elevated prenatal TH exposure accelerates early-life growth. Using cross-sectional sampling, we first report that mitochondrial DNA (mtDNA) copy number and telomere length significantly decrease from early-life to late adulthood, thus suggesting that these two molecular markers could be hallmarks of ageing in our wild bird model. Elevated prenatal THs had no effect on mtDNA copy number but counterintuitively increased telomere length both soon after birth and at the end of the growth period (equivalent to offsetting ca 4 years of post-growth telomere shortening). These findings suggest that prenatal THs might have a role in setting the 'biological' age at birth, but raise questions about the nature of the evolutionary costs of prenatal exposure to high TH levels.

Shorter telomeres precede population extinction in wild lizards.

Identifying the early warning signals of catastrophic extinctions has recently become a central focus for ecologists, but species' functional responses to environmental changes remain an untapped source for the sharpening of such warning signals. Telomere length (TL) analysis represents a promising molecular tool with which to raise the alarm regarding early population decline, since telomere attrition is associated with aging processes and accelerates after a recurrent exposure to environmental stressors. In the southern margin of their range, populations of the common lizard (Zootoca vivipara) recently became extinct at lowest elevations due to changes in climate conditions. However, the proximal signals involved in these demographic declines are still unknown. Here, we sampled 100 yearling lizards from 10 natural populations (n = 10 per population) along an extinction risk gradient. Relative lizard abundance dramatically dropped over 12 years in low-altitude populations characterized by warmer ambient temperatures and higher body growth of lizards early in life. A non-linear relationship was found between TL and population extinction risk, with shorter telomeres in populations facing high risk of extinction when compared to non-threatened ones. Our results identify TL as a promising biomarker and imply that population extinctions might be preceded by a loop of physiological aging.

Corticosterone levels in relation to trace element contamination along an urbanization gradient in the common blackbird (Turdus merula).

In a rapidly urbanizing world, trace element pollution may represent a threat to human health and wildlife, and it is therefore crucial to assess both exposition levels and associated effects of trace element contamination on urban vertebrates. In this study, we investigated the impact of urbanization on trace element contamination and stress physiology in a wild bird species, the common blackbird (Turdus merula), along an urbanization gradient (from rural to moderately urbanized areas). Specifically, we described the contamination levels of blackbirds by 4 non-essential (Ag, Cd, Hg, Pb) and 9 essential trace elements (As, Co, Cr, Cu, Fe, Mn, Ni, Se, Zn), and explored the putative disrupting effects of the non-essential element contamination on corticosterone levels (a hormonal proxy for environmental challenges). We found that non-essential trace element burden (Cd and Pb specifically) increased with increasing urbanization, indicating a significant trace element contamination even in medium sized cities and suburban areas. Interestingly, the increased feather non-essential trace element concentrations were also associated with elevated feather corticosterone levels, suggesting that urbanization probably constrains birds and that this effect may be mediated by trace element contamination. Future experimental studies are now required to disentangle the influence of multiple urban-related constraints on corticosterone levels and to specifically test the influence of each of these trace elements on corticosterone secretion.