Brian G. Walker

Field Endocrinology and Conservation Biology

Abstract Field endocrinology techniques allow the collection of samples (i.e., blood, urine, feces, tissues) from free-living animals for analysis of hormones, receptors, enzymes, etc. These data reveal mechanisms by which individuals respond to environmental challenges, breed, migrate and regulate all aspects of their life cycles. Field endocrinology techniques can also be used to address many issues in conservation biology. We briefly review past and current ways in which endocrine methods are used to monitor threatened species, identify potential stressors and record responses to environmental disturbance. We then focus on one important aspect of conservation: how free-living populations respond to human disturbance, particularly in relation to ecotourism. Breeding adult Magellanic penguins, Spheniscus magellanicus, appear to habituate well to tourists, and breed in an area where about 70,000 people visit during the season. Baseline levels of corticosterone return to normal after exposure of naïve birds to humans. However, penguin chicks appear to show a heightened adrenocortical response to handling stress in nests exposed to tourists, compared to chicks living in areas isolated from human intrusions. Given that developmental exposure to stress can have profound influences on how individuals cope with stress as adults, this potential effect of tourists on chicks could have long-term consequences. This field endocrine approach identified a stressor not observed through monitoring behavior alone.

Age and Food Deprivation Affects Expression of the Glucocorticosteroid Stress Response in Magellanic Penguin (Spheniscus magellanicus) Chicks

We examined how the glucocortical stress response in free‐living Magellanic penguin (Spheniscus magellanicus) chicks changes with age and whether adrenocortical function of chicks within a brood varies in relation to food provisioned by adults. Chicks showed little corticosterone response to capture stress shortly after hatching, an intermediate response around 45‐d posthatch, and a robust stress response near fledging. However, in response to an adrenocorticotropic hormone (ACTH) challenge, hatchlings were capable of secreting corticosterone at adult‐like levels. The larger sibling in broods of two showed a similar gradual stress‐response development pattern. In contrast, by day 45, when differences in body condition were well established between siblings, the smaller, food‐deprived chicks significantly increased baseline levels of corticosterone but showed normal stress‐induced levels. Near fledging, baseline levels had returned to normal, but stress‐induced levels were lower than expected. Similar to altricial species, normally developing semialtricial Magellanic penguin chicks do not express a robust corticosterone stress response until near fledging. Chronic stressors such as food deprivation cause corticosterone use to be up‐regulated earlier than expected. However, in cases of extended chronic stress, down‐regulation may ensue, thus avoiding the negative effects of chronically elevated levels of corticosterone.

A phylogenetically controlled test of hypotheses for behavioral insensitivity to testosterone in birds.

In most male birds that exhibit paternal care, extending the spring testosterone (T) peak throughout the breeding season reduces nestling provisioning. However, in some species, this trade-off between high T and expression of paternal care is absent. For example, during some or all of the nestling period, T did not affect paternal behavior in Male Lapland longspurs (Calcarius lapponicus), chestnut-collared longspurs (Calcarius ornatus), and great tits (Parus major). Two ecological constraints have been hypothesized to drive insensitivity to T after eggs hatch: (1) a short breeding season that limits breeding opportunities, and (2) a need for paternal care to ensure reproductive success. However, because two of the three species that exhibit T insensitivity are closely related, potential phylogenetic confounds limit determination of which, if either, factor constrains some males to T insensitivity. We examined the effects of supplementary T on paternal behavior in the Snow Bunting (Plectrophenax nivalis), a member of the monophyletic Calcarius/Plectrophenax clade. Male Snow Buntings are constrained to a short breeding season, but paternal care is not essential for survival of nestlings. We administered exogenous T during the parental phase to mimic the early spring T peak. T treatment increased song rates and interfered with paternal behavior such that nestlings of T-implanted males grew more slowly than controls. Our data suggest that T insensitivity in this clade is related to relatively recent constraints of the breeding environment (i.e., not simply common ancestry) and that the necessity of paternal care in some species may be a strong selective factor driving behavioral insensitivity to T during the parental phase.

Verification games: making verification fun

Program verification is the only way to be certain that a given piece of software is free of (certain types of) errors --- errors that could otherwise disrupt operations in the field. To date, formal verification has been done by specially-trained engineers. Labor costs have heretofore made formal verification too costly to apply beyond small, critical software components. Our goal is to make verification more cost-effective by reducing the skill set required for program verification and increasing the pool of people capable of performing program verification. Our approach is to transform the verification task (a program and a goal property) into a visual puzzle task --- a game --- that gets solved by people. The solution of the puzzle is then translated back into a proof of correctness. The puzzle is engaging and intuitive enough that ordinary people can through game-play become experts. This paper presents a status report on the Verification Games project and our Pipe Jam prototype game.

Regional Genetic Structure in the Magellanic Penguin (Spheniscus magellanicus) Suggests Metapopulation Dynamics

ABSTRACT. Using microsatellite markers and mitochondrial DNA (mtDNA) sequences (cytochrome oxidase 1 gene), we estimated levels of genetic structuring among nine Magellanic Penguin (Spheniscus magellanicus) colonies distributed throughout three major reproductive regions of the South Atlantic Ocean. Overall, breeding colonies showed relatively high levels of genetic diversity at both nuclear and mtDNA markers (mean heteorzygosity: He = 0.598; mean allelic diversity: A = 7.11; mtDNA haplotype diversity: h = 0.812). A hierarchical analysis of molecular variance based on microsatellite data showed limited genetic structuring of breeding colonies, with 99% of the variation explained by differences among individuals and 0.7–1.0% attributed to differences among the three regions. The mtDNA analysis revealed higher levels of genetic structuring, with 3.43% of the variation explained by regions and 2.24% explained by colonies within the regions. Furthermore, a Mantel test revealed a significant association between geographic and genetic distances among colonies. The limited genetic structuring we detected is likely a result of (1) population intermixing through natal dispersal and (2) the large effective sizes of the reproductive colonies, both of which prevent genetic differentiation at neutral markers, balanced with (3) the regional association of breeding colonies to distinct feeding grounds and (4) a recent expansion of the population. Our results suggest that the demographic dynamics of breeding colonies of Magellanic Penguins may be framed under a metapopulation model, in which colonies with large numbers of breeding pairs could be considered “source” populations for maintaining the overall abundance of this species in the Atlantic Ocean.

Breeding on the extreme edge: Modulation of the adrenocortical response to acute stress in two High Arctic passerines

ABSTRACT Arctic weather in spring is unpredictable and can also be extreme, so Arctic‐breeding birds must be flexible in their breeding to deal with such variability. Unpredictability in weather conditions will only intensify with climate change and this in turn could affect reproductive capability of migratory birds. Adjustments to coping strategies are therefore crucial, so here we examined the plasticity of the adrenocorticotropic stress response in two Arctic songbird species—the snow bunting (Plectrophenax nivalis) and Lapland longspur (Calcarius lapponicus)—breeding in northwest Greenland. Across the breeding season, the stress response was strongest at arrival and least robust during molt in male snow buntings. Snow bunting females had higher baseline but similar stress‐induced corticosterone levels compared to males. Modification of the stress response was not due to adrenal insensitivity, but likely regulated at the anterior pituitary gland. Compared to independent nestlings and adult snow buntings, parental‐dependent chicks had a more robust stress response. For Lapland longspurs, baseline corticosterone was highest at arrival in both male and females, and arriving males displayed a higher stress response compared to arriving females. Comparison of male corticosterone profiles collected at arrival in Greenland (76°N) and Alaska (67–71°N;) reveal that both species have higher stress responses at the more northern location. Flexibility in the stress response may be typical for birds nesting at the leading edges of their range and this ability will become more relevant as global climate change results in major shifts of breeding habitat and phenology for migratory birds. J. Exp. Zool. 323A: 266–275, 2015.

PHYSIOLOGICAL CONDITION IN MAGELLANIC PENGUINS: DOES IT MATTER IF YOU HAVE TO WALK A LONG WAY TO YOUR NEST?

Abstract Colony edges, as opposed to interiors, are often considered less advantageous nesting places in colonial species. For temperate-breeding penguins, inland colony edges should be less desirable than other edges, as there are added costs of walking farther inland, and ambient temperatures are higher. During settlement and incubation, we compared body condition and baseline and stress-induced levels of the hormone corticosterone in male Magellanic Penguins (Spheniscus magellanicus) nesting on the sea edge of a colony with those nesting on the inland edge, >800 m from shore. Body condition in both groups was significantly lower during settlement than during incubation, but was similar in both groups within breeding stages. Corticosterone levels were similar between breeding stages and for groups within each breeding stage. While body condition can vary over time, penguins appear to be well buffered to physiological extremes, as they do not show modification of corticosterone levels with variations in nesting conditions. Condiciones Fisiológicas en el Pingüino Spheniscus magellanicus: ¿Tiene Importancia Caminar una Larga Distancia al Nido? Resumen. En especies coloniales, los bordes de la colonia son generalmente considerados como lugares menos ventajosos para el establecimiento de nidos. Para los pingüinos que crían en climas templados, los bordes de las colonias orientados tierra adentro deben ser menos deseables que otros bordes, ya que se suman los costos adicionales de caminar una distancia mayor desde la orilla y debido a que las temperaturas ambientes allí son más elevadas. Durante el establecimiento y la incubación, comparamos las condiciones corporales y los niveles de referencia e inducidos por estrés de la hormona corticosterona en los machos de Spheniscus magellanicus que anidaban en el borde de la colonia adyacente al mar con los que anidaban sobre el borde que mira hacia el interior, a más de 800 m desde la orilla. La condición corporal en los dos grupos fue significantemente menor durante el establecimiento que durante la incubación, pero fue similar en los dos grupos durante las etapas de crianza. Los niveles de corticosterona fueron similares para ambas etapas de crianza y para los dos grupos durante cada etapa de crianza. Aunque la condición corporal puede variar en el tiempo, los pingüinos parecen amortiguar bien los extremos fisiológicos, ya que no muestran modificación de los niveles de corticosterona con las variaciones en las condiciones de nidificación.

Effects of El Niño and La Niña Southern Oscillation Events on the Adrenocortical Responses to Stress in Birds of the Galapagos Islands.

El Niño Southern Oscillation events (ENSO) and the subsequent opposite weather patterns in the following months and years (La Niña) have major climatic impacts, especially on oceanic habitats, affecting breeding success of both land and sea birds. We assessed corticosterone concentrations from blood samples during standardized protocols of capture, handling and restraint to simulate acute stress from 12 species of Galapagos Island birds during the ENSO year of 1998 and a La Niña year of 1999. Plasma levels of corticosterone were measured in samples collected at capture (to represent non-stressed baseline) and subsequently up to 1 h post-capture to give maximum corticosterone following acute stress, and total amount of corticosterone that the individual was exposed to during the test period (integrated corticosterone). Seabird species that feed largely offshore conformed to the brood value hypothesis whereas inshore feeding species showed less significant changes. Land birds mostly revealed no differences in the adrenocortical responses to acute stress from year to year with the exception of two small species (<18 g) that had an increase in baseline and stress responses in the ENSO year - contrary to predictions. We suggest that a number of additional variables, including body size and breeding stage may have to be considered as explanations for why patterns in some species deviated from our predictions. Nevertheless, comparative studies like ours are important for improving our understanding of the hormonal and reproductive responses of vertebrates to large scale weather patterns and global climate change in general.

Breeding on the extreme edge

ABSTRACT Arctic weather in spring is unpredictable and can also be extreme, so Arctic‐breeding birds must be flexible in their breeding to deal with such variability. Unpredictability in weather conditions will only intensify with climate change and this in turn could affect reproductive capability of migratory birds. Adjustments to coping strategies are therefore crucial, so here we examined the plasticity of the adrenocorticotropic stress response in two Arctic songbird species—the snow bunting (Plectrophenax nivalis) and Lapland longspur (Calcarius lapponicus)—breeding in northwest Greenland. Across the breeding season, the stress response was strongest at arrival and least robust during molt in male snow buntings. Snow bunting females had higher baseline but similar stress‐induced corticosterone levels compared to males. Modification of the stress response was not due to adrenal insensitivity, but likely regulated at the anterior pituitary gland. Compared to independent nestlings and adult snow buntings, parental‐dependent chicks had a more robust stress response. For Lapland longspurs, baseline corticosterone was highest at arrival in both male and females, and arriving males displayed a higher stress response compared to arriving females. Comparison of male corticosterone profiles collected at arrival in Greenland (76°N) and Alaska (67–71°N;) reveal that both species have higher stress responses at the more northern location. Flexibility in the stress response may be typical for birds nesting at the leading edges of their range and this ability will become more relevant as global climate change results in major shifts of breeding habitat and phenology for migratory birds. J. Exp. Zool. 323A: 266–275, 2015.

Breeding on the extreme edge: Modulation of the adrenocortical response to acute stress in two High Arctic passerines: STRESS RESPONSE IN TWO HIGH ARCTIC PASSERINES

ABSTRACT Arctic weather in spring is unpredictable and can also be extreme, so Arctic‐breeding birds must be flexible in their breeding to deal with such variability. Unpredictability in weather conditions will only intensify with climate change and this in turn could affect reproductive capability of migratory birds. Adjustments to coping strategies are therefore crucial, so here we examined the plasticity of the adrenocorticotropic stress response in two Arctic songbird species—the snow bunting (Plectrophenax nivalis) and Lapland longspur (Calcarius lapponicus)—breeding in northwest Greenland. Across the breeding season, the stress response was strongest at arrival and least robust during molt in male snow buntings. Snow bunting females had higher baseline but similar stress‐induced corticosterone levels compared to males. Modification of the stress response was not due to adrenal insensitivity, but likely regulated at the anterior pituitary gland. Compared to independent nestlings and adult snow buntings, parental‐dependent chicks had a more robust stress response. For Lapland longspurs, baseline corticosterone was highest at arrival in both male and females, and arriving males displayed a higher stress response compared to arriving females. Comparison of male corticosterone profiles collected at arrival in Greenland (76°N) and Alaska (67–71°N;) reveal that both species have higher stress responses at the more northern location. Flexibility in the stress response may be typical for birds nesting at the leading edges of their range and this ability will become more relevant as global climate change results in major shifts of breeding habitat and phenology for migratory birds. J. Exp. Zool. 323A: 266–275, 2015.