Seasonal Hair Glucocorticoid Fluctuations in Wild Mice (Phyllotis darwini) within a Semi-Arid Landscape in North-Central Chile.

Mammals in drylands face environmental challenges exacerbated by climate change. Currently, human activity significantly impacts these environments, and its effects on the energy demands experienced by individuals have not yet been determined. Energy demand in organisms is managed through elevations in glucocorticoid levels, which also vary with developmental and health states. Here, we assessed how anthropization, individual characteristics, and seasonality influence hair glucocorticoid concentration in the Darwin's leaf-eared mouse (Phyllotis darwini) inhabiting two areas with contrasting anthropogenic intervention in a semi-arid ecosystem of northern Chile. Hair samples were collected (n = 199) to quantify hair corticosterone concentration (HCC) using enzyme immunoassays; additionally, sex, body condition, and ectoparasite load were recorded. There were no differences in HCC between anthropized areas and areas protected from human disturbance; however, higher concentrations were recorded in females, and seasonal fluctuations were experienced by males. The results indicate that animals inhabiting semi-arid ecosystems are differentially stressed depending on their sex. Additionally, sex and season have a greater impact on corticosterone concentration than anthropogenic perturbation, possibly including temporal factors, precipitation, and primary production. The influence of sex and seasonality on HCC in P. darwini make it necessary to include these variables in future stress assessments of this species.

Associations between glucocorticoids and habitat selection reflect daily and seasonal energy requirements.

BACKGROUND: Glucocorticoids are often associated with stressful environments, but they are also thought to drive the best strategies to improve fitness in stressful environments. Glucocorticoids improve fitness in part by regulating foraging behaviours in response to daily and seasonal energy requirements. However, many studies demonstrating relationships between foraging behaviour and glucocorticoids are experimental, and few observational studies conducted under natural conditions have tested whether changing glucocorticoid levels are related to daily and seasonal changes in energy requirements. METHODS: We integrated glucocorticoids into habitat selection models to test for relationships between foraging behaviour and glucocorticoid levels in elk (Cervus canadensis) as their daily and seasonal energy requirements changed. Using integrated step selection analysis, we tested whether elevated glucocorticoid levels were related to foraging habitat selection on a daily scale and whether that relationship became stronger during lactation, one of the greatest seasonal periods of energy requirement for female mammals. RESULTS: We found stronger selection of foraging habitat by female elk with elevated glucocorticoids (eß = 1.44 95% CI 1.01, 2.04). We found no difference in overall glucocorticoid levels after calving, nor a significant change in the relationship between glucocorticoids and foraging habitat selection at the time of calving. However, we found a gradual increase in the relationship between glucocorticoids and habitat selection by female elk as their calves grew over the next few months (eß = 1.01, 95% CI 1.00, 1.02), suggesting a potentially stronger physiological effect of glucocorticoids for elk with increasing energy requirements. CONCLUSIONS: We suggest glucocorticoid-integrated habitat selection models demonstrate the role of glucocorticoids in regulating foraging responses to daily and seasonal energy requirements. Ultimately, this integration will help elucidate the implications of elevated glucocorticoids under natural conditions.

High genetic load without purging in caribou, a diverse species at risk.

High intra-specific genetic diversity is associated with adaptive potential, which is key for resilience to global change. However, high variation may also support deleterious alleles through genetic load, thereby increasing the risk of inbreeding depression if population sizes decrease. Purging of deleterious variation has been demonstrated in some threatened species. However, less is known about the costs of declines and inbreeding in species with large population sizes and high genetic diversity even though this encompasses many species globally that are expected to undergo population declines. Caribou is a species of ecological and cultural significance in North America with a wide distribution supporting extensive phenotypic variation but with some populations undergoing significant declines resulting in their at-risk status in Canada. We assessed intra-specific genetic variation, adaptive divergence, inbreeding, and genetic load across populations with different demographic histories using an annotated chromosome-scale reference genome and 66 whole-genome sequences. We found high genetic diversity and nine phylogenomic lineages across the continent with adaptive diversification of genes, but also high genetic load among lineages. We found highly divergent levels of inbreeding across individuals, including the loss of alleles by drift but not increased purging in inbred individuals, which had more homozygous deleterious alleles. We also found comparable frequencies of homozygous deleterious alleles between lineages regardless of nucleotide diversity. Thus, further inbreeding may need to be mitigated through conservation efforts. Our results highlight the "double-edged sword" of genetic diversity that may be representative of other species atrisk affected by anthropogenic activities.

An integrative and multi-indicator approach for wildlife health applied to an endangered caribou herd.

Assessing wildlife health in remote regions requires a multi-faceted approach, which commonly involves convenient samplings and the need of identifying and targeting relevant and informative indicators. We applied a novel wildlife health framework and critically assessed the value of different indicators for understanding the health status and trends of an endangered tundra caribou population. Samples and data from the Dolphin and Union caribou herd were obtained between 2015 and 2021, from community-based surveillance programs and from captured animals. We documented and categorized indicators into health determinants (infectious diseases and trace elements), processes (cortisol, pathology), and health outcomes (pregnancy and body condition). During a recent period of steep population decline, our results indicated a relatively good body condition and pregnancy rates, and decreasing levels of stress, along with a low adult cow survival. We detected multiple factors as potential contributors to the reduced survival, including Brucella suis biovar 4, Erysipelothrix rhusiopathiae and lower hair trace minerals. These results remark the need of targeted studies to improve detection and investigations on caribou mortalities. We also identified differences in health indicators between captured and hunter sampled caribou, highlighting the importance of accounting for sampling biases. This integrative approach that drew on multiple data sources has provided unprecedented knowledge on the health in this herd and highlights the value of documenting individual animal health to understand causes of wildlife declines.

Bovid Interspecies Somatic Cell Nuclear Transfer with Ooplasm Transfer.

Interspecies somatic cell nuclear transfer (iSCNT) contributes to the preservation of endangered species, albeit nuclear-mitochondrial incompatibilities constrain its application. iSCNT, coupled with ooplasm transfer (iSCNT-OT), has the potential to overcome the challenges associated with species- and genus-specific differences in nuclear-mitochondrial communication. Our iSCNT-OT protocol combines the transfer of both bison (Bison bison bison) somatic cell and oocyte ooplasm by a two-step electrofusion into bovine (Bos taurus) enucleated oocytes. The procedures described herein could be used in further studies to determine the effects of crosstalk between nuclear and ooplasmic components in embryos carrying genomes from different species.

Chromosome-level assembly of the Rangifer tarandus genome and validation of cervid and bovid evolution insights.

BACKGROUND: Genome assembly into chromosomes facilitates several analyses including cytogenetics, genomics and phylogenetics. Despite rapid development in bioinformatics, however, assembly beyond scaffolds remains challenging, especially in species without closely related well-assembled and available reference genomes. So far, four draft genomes of Rangifer tarandus (caribou or reindeer, a circumpolar distributed cervid species) have been published, but none with chromosome-level assembly. This emblematic northern species is of high interest in ecological studies and conservation since most populations are declining. RESULTS: We have designed specific probes based on Oligopaint FISH technology to upgrade the latest published reindeer and caribou chromosome-level genomes. Using this oligonucleotide-based method, we found six mis-assembled scaffolds and physically mapped 68 of the largest scaffolds representing 78% of the most recent R. tarandus genome assembly. Combining physical mapping and comparative genomics, it was possible to document chromosomal evolution among Cervidae and closely related bovids. CONCLUSIONS: Our results provide validation for the current chromosome-level genome assembly as well as resources to use chromosome banding in studies of Rangifer tarandus.

Strategies for oocyte collection and in vitro embryo production in free-roaming bison herds.

The study was conducted to test the feasibility of protocols for field collection of cumulus-oocyte complexes (COC) for in vitro embryo production (IVP) in wild bison. The study was done with captive wood bison during the anovulatory season. In Experiment 1, the efficiency of transvaginal ultrasound-guided COC collection was compared between bison restrained in a squeeze chute without sedation vs in lateral recumbency after chemical immobilization using a dart gun (n = 8/group). In Experiment 2, a 2 × 2 design was used to examine the effects of superstimulation treatment [single dose of equine chorionic gonodotrophin (eCG) vs multiple doses of follicle stimulating hormone (FSH)] and method of drug administration (manual injection vs field darting) on COC collection and IVP. In Experiment 1, no difference was detected between chute-restrained vs chemically immobilized groups in the time required to complete COC collections, the number of follicles aspirated (11.5 ± 1.9 vs 9.3 ± 1.8; P = 0.4) or the COC recovery rate [COC recovered/follicle aspirated; 58/92 (63%) vs 44/69 (64%); P = 0.9]. In Experiment 2, no differences were detected between superstimulation treatments (eCG vs FSH). The total number of follicles available for aspiration did not differ between manual injection and field darting (23.9 ± 2.7 vs 21.6 ± 1.9; P = 0.4). Compared with the random start unstimulated group, the embryo production rate was higher [18/132 (14%) vs 53/189 (28%); P = 0.04] after wave synchronization and superstimulation. Results suggest that COC collection is equally feasible in a recumbent position after chemical immobilization as those bison restrained in a standing position in a hydraulic chute. Ovarian superstimulation with a single-dose eCG protocol is as effective as a multiple-dose FSH protocol, and field darting is as effective as chute-side administration of superstimulation treatments. The strategies in the present study are ready to be incorporated into field collections in free-roaming bison herds.

How individual, social, and ecological conditions influence dispersal decisions in male vervet monkeys.

Dispersal between social groups reduces the risk of inbreeding and can improve individuals' reproductive opportunities. However, this movement has costs, such as increased risk of predation and starvation, loss of allies and kin support, and increased aggression associated with entering the new group. Dispersal strategies, such as the timing of movement and decisions on whether to transfer alone or in parallel with a peer, involve different costs and benefits. We used demographic, behavioral, hormonal, and ecological data to examine the causes and consequences of 36 dispersal events from 29 male vervet monkeys (Chlorocebus pygerythrus) at Lake Nabugabo, Uganda. Adult males' secondary dispersal coincided with the conception season in females, and males improved their potential access to females by moving to groups with higher female-to-male sex ratios and/or by increasing their dominance rank. Males that dispersed with a peer had lower fecal glucocorticoid and androgen metabolite levels than lone dispersers. Subadult males were not more likely to engage in parallel dispersals compared to adult males. Dispersal was also used as a mechanism to avoid inbreeding, but changes in hormone levels did not seem to be a trigger of dispersal in our population. Our findings illustrate the complex individual strategies used during dispersal, how many factors can influence movement decisions, as well as the value of dominance and hormone analyses for understanding these strategies.

Influence of ovarian follicular wave synchronization and single-dose eCG superstimulation on oocyte collection and in vitro embryo production in bison during the ovulatory and anovulatory seasons.

In an effort to develop an effective, minimum-handling protocol for the conservation of wood bison, the present study was designed to determine the effects of ovarian synchronization and superstimulation on cumulus oocyte complex (COC) collection and in vitro embryo production in wood bison during the ovulatory (Exp. 1) and anovulatory seasons (Exp. 2). We tested the hypotheses that COC collection and in vitro embryo production are 1) greater after follicular wave synchronization than at random stages of the follicular wave, 2) repeatable within individuals, 3) greater after ovarian superstimulation with a single dose of eCG than without treatment, and 4) greater during the anovulatory season than the ovulatory season. In Exp. 1, ultrasound-guided COC collection was performed on Day -1 in wood bison to induce follicular wave emergence the following day (Day = 0). Immediately after the COC collection on Day -1, bison were given a single im dose of 2500 IU eCG or saline (n = 6 per group). Subsequent COC collections were on Days 4 and 9. A similar design was used in Exp. 2, with an additional treatment group given 5000 IU eCG (n = 8 per group). In Exp. 1, compared to the saline-treated group, a single dose of 2500 IU eCG resulted in a greater number of ≥8 mm follicles at the time of the Day 4 COC collection (P = 0.03), but not at the Day 9. In Exp. 2, treatment with 5000 IU eCG resulted in a greater number of ≥8 mm follicles than 2500 IU eCG or the saline treatment (37.5 ± 6.9, 17.5 ± 2.0, 16.9 ± 2.0; P = 0.01, respectively). Although the number of embryos produced/COC submitted to IVM was not different among groups (mean = 18.6%), treatment with 5000 IU eCG produced more than twice as many embryos per bison as unstimulated bison (0.8 vs 1.9). In summary, embryo production rates were higher from COC collected subsequent to follicular wave synchronization vs random stages of the wave, and ovarian superstimulation with eCG resulted in a dose-related increase in the number of ≥8 mm follicles, COC collected, and embryos produced. Repeated COC collections after successive wave synchronization resulted in similar follicular counts and embryo production rates within individuals, and the greatest number of follicles aspirated, COC collected, and embryos produced was in the anovulatory season. We conclude that the minimum-handling COC collection protocols in the present study are effective and provide realistic options for embryo production in wild bison.

Qiviut cortisol is associated with metrics of health and other intrinsic and extrinsic factors in wild muskoxen (Ovibos moschatus).

Glucocorticoid (GC) levels are increasingly and widely used as biomarkers of hypothalamic-pituitary-adrenal (HPA) axis activity to study the effects of environmental changes and other perturbations on wildlife individuals and populations. However, identifying the intrinsic and extrinsic factors that influence GC levels is a key step in endocrinology studies to ensure accurate interpretation of GC responses. In muskoxen, qiviut (fine woolly undercoat hair) cortisol concentration is an integrative biomarker of HPA axis activity over the course of the hair's growth. We gathered data from 219 wild muskoxen harvested in the Canadian Arctic between October 2015 and May 2019. We examined the relationship between qiviut cortisol and various intrinsic (sex, age, body condition and incisor breakage) and extrinsic biotic factors (lungworm and gastrointestinal parasite infections and exposure to bacteria), as well as broader non-specific landscape and temporal features (geographical location, season and year). A Bayesian approach, which allows for the joint estimation of missing values in the data and model parameters estimates, was applied for the statistical analyses. The main findings include the following: (i) higher qiviut cortisol levels in males than in females; (ii) inter-annual variations; (iii) higher qiviut cortisol levels in a declining population compared to a stable population; (iv) a negative association between qiviut cortisol and marrow fat percentage; (v) a relationship between qiviut cortisol and the infection intensity of the lungworm Umingmakstrongylus pallikuukensis, which varied depending on the geographical location; and (vi) no association between qiviut cortisol and other pathogen exposure/infection intensity metrics. This study confirmed and further identified important sources of variability in qiviut cortisol levels, while providing important insights on the relationship between GC levels and pathogen exposure/infection intensity. Results support the use of qiviut cortisol as a tool to monitor temporal changes in HPA axis activity at a population level and to inform management and conservation actions.

Stress-induced changes in body surface temperature are repeatable, but do not differ between urban and rural birds.

Urbanisation can alter local microclimates, thus creating new thermal challenges for resident species. However, urban environments also present residents with frequent, novel stressors (e.g., noise, human interaction) which may demand investment in costly, self-preserving responses (e.g., the fight-or-flight response). One way that urban residents might cope with this combination of demands is by using regional heterothermy to reduce costs of thermoregulation during the stress response. In this study, we used black-capped chickadees (nurban = 9; nrural = 10) to test whether known heterothermic responses to stress exposure (here, at the bare skin around the eye): (1) varied consistently among individuals (i.e., were repeatable), and (2) were most pronounced among urban individuals compared with rural individuals. Further, to gather evidence for selection on stress-induced heterothermic responses in urban settings, we tested: (3) whether repeatability of this response was lower among birds sampled from urban environments compared with those sampled from rural environments. For the first time, we show that heterothermic responses to stress exposures (i.e. changes in body surface temperature) were highly repeatable across chronic time periods (R = 0.58) but not acute time periods (R = 0.13). However, we also show that these responses did not differ between urban and rural birds, nor were our repeatability estimates any lower in our urban sample. Thus, while regional heterothermy during stress exposure may provide energetic benefits to some, but not all, individuals, enhanced use of this response to cope with urban pressures appears unlikely in our study species.

Luteal phase length, endometrial edema, and behavior differentiate post-ovulatory events in a giant panda (Ailuropoda melanoleuca).

Despite decades of reproductive research on the giant panda (Ailuropoda melanoleuca), the post-ovulatory phase continues to confound zoologists in conservation and breeding centers around the world, often resulting in significant investments of time and resources without reproductive success. The purpose of this project was to document and compare post-ovulatory characteristics during a non-productive and productive breeding in the same individual in consecutive years. A multidisciplinary approach was used to monitor the visiting female giant panda at the Toronto Zoo through the luteal phase of her first two full reproductive cycles in 2014 and 2015. Monitoring occurred via urine-endocrine analysis, weekly ultrasound examinations, and continuous behavioral observations. The 2014 reproductive cycle consisted of a pseudopregnancy, characterized by an extended luteal phase (241 days), the identification of endometrial edema and folding during ultrasound examinations, and a lack of strong association between behavior patterns and urinary progestagen secretion. The 2015 reproductive cycle included increased feeding time through the primary progestagen rise compared to the previous year, followed by simultaneous decrease in appetite and increases in inactivity, resting, sitting upright, and pre-partum-associated behaviors. These changes began 25 days before the birth of twins on Day 153 post-ovulation. Both fetuses were detected via ultrasound 15 days pre-partum. These results suggest that an absence of pre-partum behaviors, ultrasound evidence of endometrial edema without a fetus, and an extended luteal phase may be indicative of pseudopregnancy in giant pandas. Simultaneous monitoring of morphology, behavior, and urinary-endocrine profiles showed clear differences between successful and unsuccessful reproductive years.

BPA and BPS affect the expression of anti-Mullerian hormone (AMH) and its receptor during bovine oocyte maturation and early embryo development.

BACKGROUND: Exposure to endocrine-disrupting chemicals, such as Bisphenol A (BPA) and Bisphenol S (BPS), is widespread and has negative implications on embryonic development. Preliminary evidence revealed that in women undergoing IVF treatment, urinary BPA levels were associated with low serum anti-Mullerian hormone, however a definitive relationship between the two has not yet been characterized. METHODS: This study aimed to evaluate BPA and BPS effects on in vitro oocyte maturation and early preimplantation embryo development through i) analysis of anti-Mullerian hormone (AMH) and anti-Mullerian hormone receptor II (AMHRII), ii) investigation of developmental parameters, such as cleavage, blastocyst rates and developmental arrest, iii) detection of apoptosis and iv) assessment of possible sex ratio skew. An in vitro bovine model was used as a translational model for human early embryonic development. We first assessed AMH and AMHRII levels after bisphenol exposure during oocyte maturation. Zygotes were also analyzed during cleavage and blastocysts stages. Techniques used include in vitro fertilization, quantitative polymerase chain reaction (qPCR), western blotting, TUNEL and immunofluorescence. RESULTS: Our findings show that BPA significantly decreased cleavage (p < 0.001), blastocyst (p < 0.005) and overall developmental rates as well as significantly increased embryonic arrest at the 2-4 cell stage (p < 0.05). Additionally, both BPA and BPS significantly increased DNA fragmentation in 2-4 cells, 8-16 cells and blastocyst embryos (p < 0.05). Furthermore, BPA and BPS alter AMH and AMHRII at the mRNA and protein level in both oocytes and blastocysts. BPA, but not BPS, also significantly skews sex ratios towards female blastocysts (p < 0.05). CONCLUSION: This study shows that BPA affects AMH and AMHRII expression during oocyte maturation and that BPS exerts its effects to a greater extent after fertilization and therefore may not be a safer alternative to BPA. Our data lay the foundation for future functional studies, such as receptor kinetics, downstream effectors, and promoter activation/inhibition to prove a functional relationship between bisphenols and the AMH signalling system.

Inter- and intra-specific variation in hair cortisol concentrations of Neotropical bats.

Quantifying hair cortisol has become popular in wildlife ecology for its practical advantages for evaluating stress. Before hair cortisol levels can be reliably interpreted, however, it is key to first understand the intrinsic factors explaining intra- and inter-specific variation. Bats are an ecologically diverse group of mammals that allow studying such variation. Given that many bat species are threatened or have declining populations in parts of their range, minimally invasive tools for monitoring colony health and identifying cryptic stressors are needed to efficiently direct conservation efforts. Here we describe intra- and inter-specific sources of variation in hair cortisol levels in 18 Neotropical bat species from Belize and Mexico. We found that fecundity is an important ecological trait explaining inter-specific variation in bat hair cortisol. Other ecological variables such as colony size, roost durability and basal metabolic rate did not explain hair cortisol variation among species. At the individual level, females exhibited higher hair cortisol levels than males and the effect of body mass varied among species. Overall, our findings help validate and accurately apply hair cortisol as a monitoring tool in free-ranging bats.

Analysis of hair steroid hormones in polar bears (Ursus maritimus) via liquid chromatography-tandem mass spectrometry: comparison with two immunoassays and application for longitudinal monitoring in zoos.

Analysis of hair cortisol concentrations (HCCs) is a promising method for monitoring long-term stress in mammals. However, previous measurements of HCCs in polar bears (Ursus maritimus) have yielded highly variable results, which are likely due to different methodological approaches. In this study, hair samples of zoo-housed polar bears were analyzed for cortisol with two independent immunoassays [an enzyme-linked immunoassay (EIA) and a chemiluminescence assay (CLIA)] and liquid chromatography-tandem mass spectrometry (LC-MS/MS). HCC measurements depended significantly on assay type applied, sample processing (cutting vs. powdering hair) and their interaction. Best agreement was observed between LC-MS/MS and CLIA (R2 = 0.81 for powdered hair) and sample processing had a minor, albeit significant, effect on obtained HCC measurements in these assays (R2 > 0.9). EIA measurements were consistently higher than with the other assays. HCC measurement was validated biologically for CLIA and LC-MS/MS in one male polar bear that experienced considerable stress for a prolonged period of time (> 18 weeks). Subsequently, by using the validated LC-MS/MS the measurement of cortisol could be complemented by the analysis of other steroids including cortisone, testosterone and progesterone levels from hair samples collected over a 9-month period (5-13 months) from six zoo-housed polar bears (five males, one female). No seasonal steroid variation was observed except in male progesterone levels. For all steroids except cortisone, a strong body region effect (neck or paw) was observed. Cortisol and cortisone, as well as progesterone and testosterone, concentrations were positively correlated. We show that hair steroid concentrations can be used to longitudinally measure stress and reproductive hormone axes in polar bears. The data established herein provide important basic information regarding methodology and study design for assessing hair steroid hormones.

Chronic noise exposure has context-dependent effects on stress physiology in nestling Tree Swallows (Tachycineta bicolor).

Anthropogenic noise is increasing in intensity and scope, resulting in changes to acoustic landscapes and largely negative effects on a range of species. In birds, noise can mask acoustic signals used in a variety of communication systems, including parent-offspring communication. As a result, nestling birds raised in noise may have challenges soliciting food from parents and avoiding detection by predators. Given that passerine nestlings are confined to a nest and therefore cannot escape these challenges, noise may also act as a chronic stressor during their development. Here, we raised Tree Swallow (Tachycineta bicolor) nestlings with or without continuous, white noise to test whether noise exposure affected baseline and stress-induced plasma, integrated feather corticosterone levels, and immune function. Stress physiology and immune function may also vary with the competitive environment during development, so we also examined whether noise effects varied with brood size and nestling mass. We found that overall, exposure to noise did not alter nestling stress physiology or immune function. However, light nestlings raised in noise exhibited lower baseline plasma and integrated feather corticosterone than heavy nestlings, suggesting alternative physiological responses to anthropogenic stimuli. Furthermore, light nestlings in larger broods had reduced PHA-induced immune responses compared to heavy nestlings, and PHA-induced immune responses were associated with higher levels of baseline plasma and feather CORT. Overall, our findings suggest that noise can alter the stress physiology of developing birds; however, these effects may depend on developmental conditions and the presence of other environmental stressors, such as competition for resources. Our findings may help to explain why populations are not uniformly affected by noise.

Queen Pregnancy Increases Group Estradiol Levels in Cooperatively Breeding Naked Mole-Rats.

For cooperative species, there can be great value in the synchronization of physiological states to coordinate group behavioral states. This is evident in naked mole-rats (Heterocephalus glaber), which have the most extreme form of cooperative breeding in mammals. Colonies have a single reproductive female, "the queen," and 1-3 breeding males. These breeders are supported by adult "subordinates," which are all socially suppressed into a pre-pubertal state. Subordinates cooperate in colony maintenance, defense, and alloparental care. Prior work has reported that there may be social sharing of hormones among individuals in the colony because when the queen is pregnant, subordinates of both sexes develop enlarged nipples and female subordinates can develop vaginal perforation. We sought to document the hormonal changes and mechanisms behind these observations. We found that subordinate estradiol levels were elevated during the queen's pregnancy and were correlated with queen levels. To determine if this occurs by direct hormone-sharing, where group members uptake the hormones of conspecifics through excreta or the skin, we then tested whether treating a single subordinate in the colony with estradiol would induce the same effect in other colony members. It did not, which indicates that the influence on group estradiol levels may be specific to cues from the queen. These queen cues may be behavioral in nature, as we found that queens were less aggressive during pregnancy, which prior work has suggested may relax reproductive suppression of subordinates. Yet, levels of queen aggression alone were not associated, or were weakly associated, with their colony's estradiol levels, though our sample size examining this particular relationship was low. This is suggestive that additional queen cues of reproductive status, beyond just aggression, may be relevant in influencing the subordinate hormonal change, or that the relationship between aggression and colony estradiol levels is more subtle and would need to be elucidated with a larger sample size. These results have implications for how cooperative breeders coordinate reproduction and alloparental care, and how social cues can influence individual and group physiology.

Infrared thermography as a technique to measure physiological stress in birds: Body region and image angle matter.

In vertebrates, changes in surface temperature following exposure to an acute stressor are thought to be promising indicators of the physiological stress response that may be captured noninvasively by infrared thermography. However, the efficacy of using stress-induced changes in surface temperature as indicators of physiological stress-responsiveness requires: (1) an understanding of how such responses vary across the body, (2) a magnitude of local, stress-induced thermal responses that is large enough to discriminate and quantify differences among individuals with conventional technologies, and (3) knowledge of how susceptible measurements across different body regions are to systematic error. In birds, temperature of the bare tissues surrounding the eye (the periorbital, or "eye," region) and covering the bill have each been speculated as possible predictors of stress physiological state. Using the domestic pigeon (Columba livia domestica; n = 9), we show that stress-induced changes in surface temperature are most pronounced at the bill and that thermal responses at only the bill have sufficient resolution to detect and quantify differences in responsiveness among individuals. More importantly, we show that surface temperature estimates at the eye region experience greater error due to changes in bird orientation than those at the bill. Such error concealed detection of stress-induced thermal responses at the eye region. Our results highlight that: (1) in some species, bill temperature may serve as a more robust indicator of autonomic stress-responsiveness than eye region temperature, and (2) future studies should account for spatial orientation of study individuals if inference is to be drawn from infrared thermographic images.

Fecal adrenal hormone patterns during ovulatory and non-ovulatory reproductive cycles in female veiled chameleons (Chamaeleo calyptratus).

The relationship between the reproductive (hypothalamic-pituitary-gonadal; HPG) and adrenal (hypothalamic-pituitary-adrenal; HPA) hormone axes is complex and can vary depending on the species and environmental factors affecting an individual. In an effort to understand this relationship in female veiled chameleons (Chamaeleo calyptratus), the patterns of fecal metabolites of corticosterone (C), estradiol (E), testosterone (T), and progesterone (P) were analyzed by enzyme immunoassay (EIA) during ovulatory (OC; eggs laid) and non-ovulatory cycles (NOC; no eggs laid). Glucocorticoid (GC) metabolites in the fecal extracts were characterized by HPLC and corticosterone EIA performance was assessed by parallelism, accuracy and precision tests. The results indicated that the assay chosen reliably measured the hormone metabolites present in the fecal extracts. Regular, cyclical hormone metabolite patterns consisting of an E peak followed by peaks of T, P and C in close succession were observed during both ovulatory and non-ovulatory cycles; relative levels of P and C, however, were higher during ovulatory cycles. Corticosterone metabolite levels, in particular, increased throughout vitellogenesis and peaked in late vitellogenesis (in non-ovulatory cycles) or around the time of ovulation, and remained elevated throughout the gravid period, falling just prior to oviposition. The results provide evidence of variation in glucocorticoid production throughout different stages of the reproductive cycle, including a role in the ovulatory process; the physiology, however, remains unclear.